Methods of treating and preventing graft versus host disease

ABSTRACT

Described herein are methods for treating and preventing graft versus host disease using ACK inhibitors. The methods include administering to an individual in need thereof an ACK inhibitor such as ibrutinib for treating and preventing graft versus host disease.

CROSS-REFERENCE

This application is a divisional of U.S. application Ser. No.15/586,058, filed May 3, 2017, which is a continuation of U.S.application Ser. No. 14/823,650, filed Oct. 24, 2014, now U.S. Pat. No.9,795,604, issued Oct. 24, 2017, which claims the benefit of priority ofU.S. Provisional Application No. 61/895,981, filed Oct. 25, 2013; U.S.Provisional Application No. 61/910,945, filed Dec. 2, 2013; U.S.Provisional Application No. 61/973,173, filed Mar. 31, 2014; and U.S.Provisional Application No. 61/973,176 filed Mar. 31, 2014, each ofwhich are hereby incorporated by reference in their entireties as iffully set forth herein.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted in ASCII format via EFS-Web and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Oct. 25, 2017, isnamed PIR-88502_Sequence_Listing and is 674 bytes in size.

BACKGROUND OF THE INVENTION

Chronic graft versus host disease (cGVHD) is the most common long-termcomplication following allogeneic stem cell transplant (SCT), affecting30-70% of patients who survive beyond the first 100 days. cGVHD and itsassociated immune deficiency have been identified as a leading cause ofnon-relapse mortality (NRM) in allogeneic SCT survivors. SCT survivorswith cGVHD are 4.7 times as likely to develop severe or life-threateninghealth conditions compared with healthy siblings, and patients withactive cGVHD are more likely to report adverse general health, mentalhealth, functional impairments, activity limitation, and pain thanallo-SCT survivors with no history of cGVHD. Any organ system can beaffected, and further morbidity is frequently caused by long-termexposure to the corticosteroids and calcineurin inhibitors required totreat the condition.

SUMMARY OF THE INVENTION

Disclosed herein, in some embodiments, are methods of preventing theoccurrence of graft versus host disease (GVHD) or reducing the severityof GVHD occurrence in a patient requiring cell transplantationcomprising administration of a therapeutically effective amount of anACK inhibitor (e.g., an ITK or BTK inhibitor). In some embodiments,disclosed herein are methods of reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administration ofa therapeutically effective amount of an ACK inhibitor (e.g., an ITK orBTK inhibitor). In some embodiments the ACK inhibitor is a compound ofFormula (A). In some embodiments, disclosed herein are methods ofpreventing the occurrence of graft versus host disease (GVHD) orreducing the severity of GVHD occurrence in a patient requiring celltransplantation, comprising administration of a therapeuticallyeffective amount of a compound of Formula (A) having the structure:

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NR₉—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—,—S(═O)₂NH—, —NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—,—OC(O)NR₉—, —NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-,aryl-, —NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, −OC(═NR₁₁)—,or —C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or

two R₁₀ groups can together form a 5-, 6-, 7-, or 8-memberedheterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof. Insome embodiments, L₃, X and L₄ taken together form a nitrogen containingheterocyclic ring. In some embodiments, the nitrogen containingheterocyclic ring is a piperidine group. In some embodiments, G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3-[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematological malignancy. In some embodiments, the patienthas a relapsed or refractory hematological malignancy. In someembodiments, the patient has a B-cell malignancy. In some embodiments,the patient has a T-cell malignancy. In some embodiments, the patienthas a leukemia, a lymphoma, or a myeloma. In some embodiments, theB-cell malignancy is a non-Hodgkin's lymphoma. In some embodiments, theB-cell malignancy is chronic lymphocytic leukemia (CLL). In someembodiments, the B-cell malignancy is a relapsed or refractory B-cellmalignancy. In some embodiments, the B-cell malignancy is a relapsed orrefractory non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is a relapsed or refractory CLL. In some embodiments, thepatient has high risk CLL. In some embodiments, the patient has a 17pchromosomal deletion. In some embodiments, the patient has 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLL as determined by bonemarrow biopsy. In some embodiments, the patient has received one or moreprior anticancer agents. In some embodiments, the anticancer agent isselected from among alemtuzumab, bendamustine, bortezomib, CAL-101,chlorambucil, cyclophosphamide, dexamethasone, docetaxel, doxorubicin,endostatineverolimus, etoposide, fludarabine, fostamatinib,hydroxydaunorubicin, ibritumomab, ifosphamide, lenalidomide, mesalazine,ofatumumab, paclitaxel, pentostatin, prednisone, rituximab,temsirolimus, thalidomide, tositumomab, vincristine, or a combinationthereof. In some embodiments, the anticancer agent is rituximab. In someembodiments, the anticancer agent is alemtuzumab. In some embodiments,the anticancer agent is fludarabine, cyclophosphamide, and rituximab(FCR). In some embodiments, the anticancer agent is oxaliplatin,fludarabine, cytarabine, rituximab (OFAR). In some embodiments, theamount of the ACK inhibitor compound (e.g., a compound of Formula (A))prevents or reduces GVHD while maintaining a graft-versus-leukemia (GVL)reaction effective to reduce or eliminate the number of cancerous cellsin the blood of the patient. In some embodiments, the celltransplantation is a hematopoietic cell transplantation. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Insome embodiments, the GVHD is steroid resistant GVHD. In someembodiments, the GVHD is cyclosporin-resistant GVHD. In someembodiments, the GVHD is refractory GVHD. In some embodiments, the GHVDis oral GVHD. In some embodiments, the oral GVHD is reticular oral GVHD.In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered concurrently with an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered subsequent to an allogeneicbone marrow or hematopoietic stem cell transplant. In some embodiments,the patient is a candidate for receiving HLA-mismatched hematopoieticstem cells. In some embodiments, the patient is a candidate forreceiving unrelated donor hematopoietic stem cells, umbilical veinhematopoietic stem cells, or peripheral blood stem cells. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered orally. In some embodiments, the ACK inhibitorcompound (e.g., a compound of Formula (A)) is administered at a dosageof between about 0.1 mg/kg per day to about 100 mg/kg per day. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered at a dosage of about 40 mg/day, about 140 mg/day,about 280 mg/day, about 420 mg/day, about 560 mg/day, or about 840mg/day. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered in combination with otherprophylactic agents. In some embodiments, the ACK inhibitor compound(e.g., a compound of Formula (A)) is administered from day 1 to aboutday 120 following allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered from day 1 to about day 1000following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered in combination with one or more additionaltherapeutic agents. In some embodiments, the additional therapeuticagent is a corticosteroid. In some embodiments, the therapeutic agent iscyclosporine (CSA), mycophenolate mofetil (MMF) or a combinationthereof. In some embodiments, the patient has or will receive a donorlymphocyte infusions (DLI). In some embodiments, the patient isadministered one or more DLIs. In some embodiments, the patient isadministered two or more DLIs. In some embodiments, the DLI comprisesCD3+ lymphocytes. In some embodiments, the patient is administered oneor more donor lymphocyte infusions (DLI) following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, theACK inhibitor compound (e.g., a compound of Formula (A)) is administeredconcurrently with a DLI following allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to a DLI following an allogeneic bone marrow or hematopoietic stemcell transplant. In some embodiments, the ACK inhibitor compound (e.g.,a compound of Formula (A)) is administered following a DLI following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is ibrutinib.

Disclosed herein, in some embodiments, are methods of treating a patientfor alleviation of a bone marrow mediated disease, comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, and a therapeutically effective amount of an ACKinhibitor (e.g., an ITK or BTK inhibitor). In some embodiments,disclosed herein are methods of treating a patient for alleviation of abone marrow mediated disease, with alleviation of consequently developedgraft versus host disease (GVHD), comprising administering to thepatient allogeneic hematopoietic stem cells and/or allogeneic T-cells,and a therapeutically effective amount of a compound of Formula (A):

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—, —S(═O)₂NH—,—NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—,—NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-, aryl-,—NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or—C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or

two R₁₀ groups can together form a 5-, 6-, 7-, or 8-memberedheterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or

each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof, isadministered prior to, concurrently with, or following the allogeneichematopoietic stem cells and/or allogeneic T-cells. In some embodiments,L₃, X and L₄ taken together form a nitrogen containing heterocyclicring. In some embodiments, the nitrogen containing heterocyclic ring isa piperidine group. In some embodiments, G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3-[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematological malignancy. In some embodiments, the patienthas a relapsed or refractory hematological malignancy. In someembodiments, the patient has a leukemia, a lymphoma, or a myeloma. Insome embodiments, the patient has a B-cell malignancy. In someembodiments, the B-cell malignancy is a non-Hodgkin's lymphoma. In someembodiments, the B-cell malignancy is chronic lymphocytic leukemia(CLL). In some embodiments, the B-cell malignancy is a relapsed orrefractory B-cell malignancy. In some embodiments, the B-cell malignancyis a relapsed or refractory non-Hodgkin's lymphoma. In some embodiments,the B-cell malignancy is a relapsed or refractory CLL. In someembodiments, the patient has high risk CLL. In some embodiments, thepatient has a 17p chromosomal deletion. In some embodiments, the patienthas 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLL asdetermined by bone marrow biopsy. In some embodiments, the patient hasreceived one or more prior anticancer agents. In some embodiments, theanticancer agent is selected from among alemtuzumab, bendamustine,bortezomib, CAL-101, chlorambucil, cyclophosphamide, dexamethasone,docetaxel, doxorubicin, endostatineverolimus, etoposide, fludarabine,fostamatinib, hydroxydaunorubicin, ibritumomab, ifosphamide,lenalidomide, mesalazine, ofatumumab, paclitaxel, pentostatin,prednisone, rituximab, temsirolimus, thalidomide, tositumomab,vincristine, or a combination thereof. In some embodiments, theanticancer agent is rituximab. In some embodiments, the anticancer agentis alemtuzumab. In some embodiments, the anticancer agent isfludarabine, cyclophosphamide, and rituximab (FCR). In some embodiments,the anticancer agent is oxaliplatin, fludarabine, cytarabine, rituximab(OFAR). In some embodiments, the amount of the ACK inhibitor compound(e.g., a compound of Formula (A)) prevents or reduces GVHD whilemaintaining a graft-versus-leukemia (GVL) reaction effective to reduceor eliminate the number of cancerous cells in the blood of the patient.In some embodiments, the cell transplantation is a hematopoietic celltransplantation. In some embodiments, the GVHD is acute GVHD. In someembodiments, the GVHD is chronic GVHD. In some embodiments, the GVHD issclerodermatous GVHD. In some embodiments, the GVHD is steroid resistantGVHD. In some embodiments, the GVHD is cyclosporin-resistant GVHD. Insome embodiments, the GVHD is refractory GVHD. In some embodiments, theGHVD is oral GVHD. In some embodiments, the oral GVHD is reticular oralGVHD. In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered concurrently with an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered subsequent to an allogeneicbone marrow or hematopoietic stem cell transplant. In some embodiments,the patient is a candidate for receiving HLA-mismatched hematopoieticstem cells. In some embodiments, the patient is a candidate forreceiving unrelated donor hematopoietic stem cells, umbilical veinhematopoietic stem cells, or peripheral blood stem cells. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered at a dosage of between about 0.1 mg/kg per day toabout 100 mg/kg per day. In some embodiments, the ACK inhibitor compound(e.g., a compound of Formula (A)) is administered at a dosage of about40 mg/day, about 140 mg/day, about 280 mg/day, about 420 mg/day, about560 mg/day, or about 840 mg/day. In some embodiments, the ACK inhibitorcompound (e.g., a compound of Formula (A)) is administered orally. Insome embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered in combination with additional therapeuticagents. In some embodiments, the additional therapeutic agent is acorticosteroid. In some embodiments, the additional therapeutic agent iscyclosporine (CSA), mycophenolate mofetil (MMF) or a combinationthereof. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered from day 1 to about day 120following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered from day 1 to about day 1000 followingallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the patient has or will receive a donor lymphocyte infusion(DLI). In some embodiments, the patient has or will receive two or moredonor lymphocyte infusions (DLI). In some embodiments, the patient isadministered one or more donor lymphocyte infusions (DLI). In someembodiments, the DLI comprises CD3+ lymphocytes. In some embodiments,the patient is administered one or more donor lymphocyte infusions (DLI)following an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered concurrently with a DLIfollowing allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered prior to a DLI following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, theACK inhibitor compound (e.g., a compound of Formula (A)) is administeredfollowing a DLI following an allogeneic bone marrow or hematopoieticstem cell transplant. In some embodiments, the ACK inhibitor compound(e.g., a compound of Formula (A)) is ibrutinib.

In some embodiments, disclosed herein are methods of reducing theseverity of GVHD occurrence in a patient requiring cell transplantationcomprising administration of a therapeutically effective amount ofibrutinib(1-[(3R)-3[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one).In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematological malignancy. In some embodiments, the patienthas a relapsed or refractory hematological malignancy. In someembodiments, the patient has a B-cell malignancy. In some embodiments,the patient has a T-cell malignancy. In some embodiments, the patienthas a leukemia, a lymphoma, or a myeloma. In some embodiments, theB-cell malignancy is a non-Hodgkin's lymphoma. In some embodiments, theB-cell malignancy is chronic lymphocytic leukemia (CLL). In someembodiments, the B-cell malignancy is a relapsed or refractory B-cellmalignancy. In some embodiments, the B-cell malignancy is a relapsed orrefractory non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is a relapsed or refractory CLL. In some embodiments, thepatient has high risk CLL. In some embodiments, the patient has a 17pchromosomal deletion. In some embodiments, the patient has 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLL as determined by bonemarrow biopsy. In some embodiments, the patient has received one or moreprior anticancer agents. In some embodiments, the anticancer agent isselected from among alemtuzumab, bendamustine, bortezomib, CAL-101,chlorambucil, cyclophosphamide, dexamethasone, docetaxel, doxorubicin,endostatineverolimus, etoposide, fludarabine, fostamatinib,hydroxydaunorubicin, ibritumomab, ifosphamide, lenalidomide, mesalazine,ofatumumab, paclitaxel, pentostatin, prednisone, rituximab,temsirolimus, thalidomide, tositumomab, vincristine, or a combinationthereof. In some embodiments, the anticancer agent is rituximab. In someembodiments, the anticancer agent is alemtuzumab. In some embodiments,the anticancer agent is fludarabine, cyclophosphamide, and rituximab(FCR). In some embodiments, the anticancer agent is oxaliplatin,fludarabine, cytarabine, rituximab (OFAR). In some embodiments, theamount of ibrutinib prevents or reduces GVHD while maintaining agraft-versus-leukemia (GVL) reaction effective to reduce or eliminatethe number of cancerous cells in the blood of the patient. In someembodiments, the cell transplantation is a hematopoietic celltransplantation. In some embodiments, the GVHD is acute GVHD. In someembodiments, the GVHD is chronic GVHD. In some embodiments, the GVHD issclerodermatous GVHD. In some embodiments, the GVHD is steroid resistantGVHD. In some embodiments, the GVHD is cyclosporin-resistant GVHD. Insome embodiments, the GVHD is refractory GVHD. In some embodiments, theGHVD is oral GVHD. In some embodiments, the oral GVHD is reticular oralGVHD. In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ibrutinib is administered concurrently with anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ibrutinib is administered prior to an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, theibrutinib is administered subsequent to an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the patient isa candidate for receiving HLA-mismatched hematopoietic stem cells. Insome embodiments, the patient is a candidate for receiving unrelateddonor hematopoietic stem cells, umbilical vein hematopoietic stem cells,or peripheral blood stem cells. In some embodiments, the ibrutinib isadministered orally. In some embodiments, the ibrutinib is administeredat a dosage of between about 0.1 mg/kg per day to about 100 mg/kg perday. In some embodiments, the ibrutinib is administered at a dosage ofabout 40 mg/day, about 140 mg/day, about 280 mg/day, about 420 mg/day,about 560 mg/day, or about 840 mg/day. In some embodiments, theibrutinib is administered in combination with other prophylactic agents.In some embodiments, the ibrutinib is administered from day 1 to aboutday 120 following allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ibrutinib is administered from day1 to about day 1000 following allogeneic bone marrow or hematopoieticstem cell transplant. In some embodiments, the ibrutinib is administeredin combination with one or more additional therapeutic agents. In someembodiments, the additional therapeutic agent is a corticosteroid. Insome embodiments, the therapeutic agent is cyclosporine (CSA),mycophenolate mofetil (MMF) or a combination thereof. In someembodiments, the patient has or will receive a donor lymphocyteinfusions (DLI). In some embodiments, the patient is administered one ormore DLIs. In some embodiments, the patient is administered two or moreDLIs. In some embodiments, the DLI comprises CD3+ lymphocytes. In someembodiments, the patient is administered one or more donor lymphocyteinfusions (DLI) following an allogeneic bone marrow or hematopoieticstem cell transplant. In some embodiments, the ibrutinib is administeredconcurrently with a DLI following allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ibrutinibis administered prior to a DLI following an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ibrutinibis administered following a DLI following an allogeneic bone marrow orhematopoietic stem cell transplant.

INCORPORATION BY REFERENCE

All publications, patents, and patent applications mentioned in thisspecification are herein incorporated by reference to the same extent asif each individual publication, patent, or patent application wasspecifically and individually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

FIG. 1 exemplifies that ibrutinib ameliorates cGVHD symptomatology afterallotransplant. C57BL/6 mice were engrafted with LP/J bone marrow after850 cGy lethal irradiation. 25 days post-transplant mice were randomlyassigned to ibrutinib, vehicle, or cyclosporine groups. Panel A showsimages showing external signs of cGVHD including alopecia, scleroderma,and fibrotic lesions at day 36 post-transplant. Ibrutinib treatmentgroup displayed few external signs of cGVHD progression as compared tovehicle or cyclosporine groups. Panel B shows an analysis of cGVHD mousegroups using a physical scoring system adapted from Cooke et al., whichincorporates weight, posture, coat condition, skin condition, andmobility. Scoring was conducted on day 36 post-transplantation. Panel Cshows the LP/J→C57BL/6 cGVHD scoring. Each category: coat condition,skin condition, weight, posture, mobility, and vitality are individuallyscored and summed to achieve an overall cGVHD condition score. Scoresare taken by a consistent unbiased observer with no knowledge oftreatment cohorts. Panel D provides images of cGVHD mouse groups at day39 post-HSCT. Panel E provides images of H&E stained skin preparationsof sclerodermatous skin lesions showing levels of dermal fibrosis,epidermal hyperplasia, serocellular crusting, erosion, andlymphohistiocytic infiltration, consistent with cGVHD.

FIG. 2 exemplifies that Tregs are not inhibited by ibrutinib. Panel Aprovides a plot of FoxP3+ CD4+ cells in C57BL/6 mice treated withibrutinib (25 mg/kg/day) or vehicle for 9 weeks. The percent FoxP3+ CD4+cells was analyzed by flow cytometry on peripheral blood. Student'sT-test indicates no significant difference between the two groups. PanelB provides a plot of CD8 T cell division index for variousresponder:suppressor ratios. Purified CD4+ CD25hiCD127dim CD49d-FoxP3+Tregs were pretreated with 1 μM ibrutinib or vehicle and mixed with CFSE-labeled autologous CD8+ responder cells at the indicated ratio.Anti-CD3/CD28/CD2 stimulation beads were added and stimulation wasassessed by CFSE dilution calculated division index after 6 days.Negative control wells contained no stimulation beads. n=7; error bars:s.e.m.

FIG. 3 exemplifies that Th2 immunity was inhibited by ibrutinib. Panel Aprovides a plot of normalized intracellular staining analysis of IL4(open bars n=6) and IFNγ (closed bars n=9) CD4+ cells derived pretreatedwith ibrutinib and stimulated with anti-CD3/anti-CD28. Error bars=s.e.m.Panel B provides a plot of plasma IgG1 (Th2) and IgG2c (Th1) subisotypeanalysis of C57BL/6 EμTCL1 mice at 8 months of age after 7 consecutivemonths of ibrutinib (25 mg/kg/day) (n=12) or vehicle (n=13)administration via drinking.

FIG. 4 exemplifies that Th17 immunity was inhibited by ibrutinib. Th17cells were magnetically enriched from freshly isolated healthy donorPBMCs using CXCR3-CD4+CCL6+ isolation. After enrichment cells weretreated with ibrutinib or vehicle for 30 minutes prior to drug washout.Cells were stimulated with anti-CD3 and anti-CD28 for 12 hours withGOLGISTOP protein transport inhibitor. IL17 producing cells werequantified as a percentage of total live CD4+ T-cells and finalpercentages were normalized to DMSO group. n=3; error bars: s.e.m.

FIG. 5 exemplifies that ibrutinib inhibited cGVHD autoimmunesymptomatology and progression. Panel A provides a plot of the weeklyblinded analysis of cGVHD external metrics including weight, posture,vitality, mobility, coat, and skin. All cGVHD scores were corrected forindividual scores at the beginning of treatment (day 25). Panel Bprovides a Kaplan Meier plot of cGVHD progression free survival.Progression is defined as >2 point increase in day 25 cGVHD score.*=p<0.01 Error bars=s.e.m.

FIG. 6 exemplifies that ibrutinib therapy combats autoimmuneinfiltration of internal organs in a T-cell dependent model of cGVHD.Panel A shows representative 20× images from H&E, B220, or CD3 stainedlung and kidney tissues from mice sacrificed at day 125 post-HSCT.Images were taken by a trained veterinary pathologist who was blinded toanimal cohorts. Panel B shows a blinded pathologic analysis of H&Estained lung tissues obtained from cGVHD cohorts. Lymphohistiocyticinfiltration was graded on a 0-4 scale for each animal. Panel C shows ablinded pathologic analysis of H&E stained liver tissues obtained fromcGVHD cohorts. Portal hepatitis and vasculitis was graded on a 0-4 scalefor each animal. Panel D shows a Kaplan-Meier plot of cGVHDprogression-free survival in an independent experiment aimed todetermine sustained benefits from continued ibrutinib therapy. Duringthe course of the experiment, ibrutinib was withdrawn on day 60 fromanimals in the Ibrutinib (day 25 to day 60) cohort. **P<0.001.

FIG. 7 exemplifies that ibrutinib limits activation of T-cells andB-cells from patients with active cGVHD. Primary CD4+ T-cells wereisolated from patients with active cGVHD, pretreated with 1 μM ibrutinib(or DMSO), and stimulated using anti-CD3 for 6 hours. Panel A shows agraph depicting CD69+ CD4+ T-cell percentage for each patient. “*”indicates p<0.05. Panel B shows an image of an immunoblot analysis ofBTK, ERK, and PLCγ2 in B-cells isolated from patients with cGVHD werepretreated with 1 μM ibrutinib (or DMSO), and stimulated using anti-IgMfor 45 minutes. Data are representative of three experiments on threeseparate patients.

FIG. 8 exemplifies a clinical study of ibrutinib (PCI-32765) treatmentof a post-allogeneic HCT transplant patient with refractory CLL withoropharyngeal chronic GVHD. CLL minimum residual disease (MRD) and bloodCD3+ T cell donor chimerism is shown over time following allo-HCTtransplantation. Donor lymphocyte infusions (DLI) and initiation ofibrutinib treatment are indicated (see Example 5 for exemplary treatmentprotocol).

FIG. 9 Panels A-E shows plots depicting percent change in absolutelymphocytic count (ALC) for 2 patients who received ibrutinib treatmentfor >1 yr. SPN=Stanford Patient Number (Panel A); percent reduction inLN size, as reported by the sum of the product of LN diameters (SPD) for4 patients following initiation of ibrutinib (Panel B); CLL MRD(reported as a percentage of WBCs) and blood donor CD3 T cell levelsshown for patient SPN 3975(Panel C); B cells (excluding the CLL clone)as percent of total PBMC for patient SPN 3975 as measured by IgH HTS(Panel D); Total IgH molecules and unique IgH clone counts for patientSPN 3975 at different time points (D=day) post allo-HCT (Panel E).

DETAILED DESCRIPTION OF THE INVENTION

In some embodiments, disclosed herein is a method of preventing theoccurrence of graft versus host disease (GVHD) or reducing the severityof GVHD occurrence in a patient requiring cell transplantation,comprising administration of an ACK inhibitor compound (e.g., an ITK orBTK inhibitor compound). In some embodiments, disclosed herein is amethod of preventing the occurrence of graft versus host disease (GVHD)or reducing the severity of GVHD occurrence in a patient requiring celltransplantation, comprising administration of a therapeuticallyeffective amount of a compound of Formula (A) having the structure:

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—, —S(═O)₂NH—,—NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—,—NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-, aryl-,—NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or—C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or

two R₁₀ groups can together form a 5-, 6-, 7-, or 8-memberedheterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof. Insome embodiments, L₃, X and L₄ taken together form a nitrogen containingheterocyclic ring. In some embodiments, the nitrogen containingheterocyclic ring is a piperidine group. In some embodiments, G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematologic malignancy. In some embodiments, the patienthas a B-cell malignancy. In some embodiments, the patient has a T-cellmalignancy. In some embodiments, the patient has a leukemia, a lymphoma,or a myeloma. In some embodiments, the patient has a B-cell malignancy.In some embodiments, the B-cell malignancy is a non-Hodgkin's lymphoma.In some embodiments, the B-cell malignancy is chronic lymphocyticleukemia (CLL). In some embodiments, the B-cell malignancy is a relapsedor refractory B-cell malignancy. In some embodiments, the B-cellmalignancy is a relapsed or refractory non-Hodgkin's lymphoma. In someembodiments, the B-cell malignancy is a relapsed or refractory CLL. Insome embodiments, the patient has high risk CLL. In some embodiments,the patient has a 17p chromosomal deletion. In some embodiments, thepatient has 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLLas determined by bone marrow biopsy. In some embodiments, the patienthas received one or more prior anticancer agents. In some embodiments,the anticancer agent is selected from among alemtuzumab, bendamustine,bortezomib, CAL-101, chlorambucil, cyclophosphamide, dexamethasone,docetaxel, doxorubicin, endostatineverolimus, etoposide, fludarabine,fostamatinib, hydroxydaunorubicin, ibritumomab, ifosphamide,lenalidomide, mesalazine, ofatumumab, paclitaxel, pentostatin,prednisone, rituximab, temsirolimus, thalidomide, tositumomab,vincristine, or a combination thereof. In some embodiments, theanticancer agent is rituximab. In some embodiments, the anticancer agentis alemtuzumab. In some embodiments, the anticancer agent isfludarabine, cyclophosphamide, and rituximab (FCR). In some embodiments,the anticancer agent is oxaliplatin, fludarabine, cytarabine, rituximab(OFAR). In some embodiments, the amount of the compound of Formula (A)prevents or reduces GVHD while maintaining a graft-versus-leukemia (GVL)reaction effective to reduce or eliminate the number of cancerous cellsin the blood of the patient. In some embodiments, the celltransplantation is a hematopoietic cell transplantation. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Insome embodiments, the GVHD is steroid resistant GVHD. In someembodiments, the GVHD is cyclosporin-resistant GVHD. In someembodiments, the GVHD is refractory GVHD. In some embodiments, the GHVDis oral GVHD. In some embodiments, the oral GVHD is reticular oral GVHD.In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the compound of Formula (A) is administered concurrentlywith an allogeneic bone marrow or hematopoietic stem cell transplant. Insome embodiments, the compound of Formula (A) is administered prior toan allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the compound of Formula (A) is administered following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the patient is a candidate for receiving HLA-mismatchedhematopoietic stem cells. In some embodiments, the patient is acandidate for receiving unrelated donor hematopoietic stem cells,umbilical vein hematopoietic stem cells, or peripheral blood stem cells.In some embodiments, the compound of Formula (A) is administered at adosage of between about 0.1 mg/kg per day to about 100 mg/kg per day. Insome embodiments, the compound of Formula (A) is administered at adosage of about 40 mg/day, about 140 mg/day, about 280 mg/day, about 420mg/day, about 560 mg/day, or about 840 mg/day. In some embodiments, thecompound of Formula (A) is administered in combination with additionaltherapeutic agents. In some embodiments, the additional therapeuticagent is a corticosteroid. In some embodiments, the additionaltherapeutic agent is cyclosporine (CSA), mycophenolate mofetil (MMF) ora combination thereof. In some embodiments, the compound of Formula (A)is administered orally. In some embodiments, the compound of Formula (A)is administered from day 1 to about day 120 following allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, thecompound of Formula (A) is administered from day 1 to about day 1000following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the patient is administered one or more donorlymphocyte infusions (DLI). In some embodiments, the DLI comprises CD3+lymphocytes. In some embodiments, the patient is administered one ormore donor lymphocyte infusions (DLI) following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, thecompound of Formula (A) is administered concurrently with a DLIfollowing allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the compound of Formula (A) is administered priorto a DLI following an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the compound of Formula (A) isadministered following a DLI following an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the compound ofFormula (A) is ibrutinib.

Disclosed herein, in some embodiments, is a method of treating a patientfor alleviation of a bone marrow mediated disease, with alleviation ofconsequently developed graft versus host disease (GVHD), comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, wherein a therapeutically effective amount of acompound of Formula (A):

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NR₉—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—,—S(═O)₂NH—, —NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—,—OC(O)NR₉—, —NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-,aryl-, —NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—,or —C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or two R₁₀ groups cantogether form a 5-, 6-, 7-, or 8-membered heterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof, isadministered prior to, concurrently with, or following theadministration allogeneic hematopoietic stem cells and/or allogeneicT-cells. In some embodiments, L₃, X and L₄ taken together form anitrogen containing heterocyclic ring. In some embodiments, the nitrogencontaining heterocyclic ring is a piperidine group. In some embodiments,G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3-[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient as a hematologic malignancy. In some embodiments, the patienthas a B-cell malignancy. In some embodiments, the patient has a T-cellmalignancy. In some embodiments, the patient has a leukemia, a lymphoma,or a myeloma. In some embodiments, the patient has a B-cell malignancy.In some embodiments, the B-cell malignancy is a non-Hodgkin's lymphoma.In some embodiments, the B-cell malignancy is chronic lymphocyticleukemia (CLL). In some embodiments, the B-cell malignancy is a relapsedor refractory B-cell malignancy. In some embodiments, the B-cellmalignancy is a relapsed or refractory non-Hodgkin's lymphoma. In someembodiments, the B-cell malignancy is a relapsed or refractory CLL. Insome embodiments, the patient has high risk CLL. In some embodiments,the patient has a 17p chromosomal deletion. In some embodiments, thepatient has 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLLas determined by bone marrow biopsy. In some embodiments, the patienthas received one or more prior anticancer agents. In some embodiments,the anticancer agent is selected from among alemtuzumab, bendamustine,bortezomib, CAL-101, chlorambucil, cyclophosphamide, dexamethasone,docetaxel, doxorubicin, endostatineverolimus, etoposide, fludarabine,fostamatinib, hydroxydaunorubicin, ibritumomab, ifosphamide,lenalidomide, mesalazine, ofatumumab, paclitaxel, pentostatin,prednisone, rituximab, temsirolimus, thalidomide, tositumomab,vincristine, or a combination thereof. In some embodiments, theanticancer agent is rituximab. In some embodiments, the anticancer agentis alemtuzumab. In some embodiments, the anticancer agent isfludarabine, cyclophosphamide, and rituximab (FCR). In some embodiments,the anticancer agent is oxaliplatin, fludarabine, cytarabine, rituximab(OFAR). In some embodiments, the amount of the compound of Formula (A)prevents or reduces GVHD while maintaining a graft-versus-leukemia (GVL)reaction effective to reduce or eliminate the number of cancerous cellsin the blood of the patient. In some embodiments, the celltransplantation is a hematopoietic cell transplantation. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Insome embodiments, the GVHD is steroid resistant GVHD. In someembodiments, the GVHD is cyclosporin-resistant GVHD. In someembodiments, the GVHD is refractory GVHD. In some embodiments, the GHVDis oral GVHD. In some embodiments, the oral GVHD is reticular oral GVHD.In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the compound of Formula (A) is administered concurrentlywith an allogeneic bone marrow or hematopoietic stem cell transplant. Insome embodiments, the compound of Formula (A) is administered prior toan allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the compound of Formula (A) is administered following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the patient is a candidate for receiving HLA-mismatchedhematopoietic stem cells. In some embodiments, the patient is acandidate for receiving unrelated donor hematopoietic stem cells,umbilical vein hematopoietic stem cells, or peripheral blood stem cells.In some embodiments, the compound of Formula (A) is administered at adosage of between about 0.1 mg/kg per day to about 100 mg/kg per day. Insome embodiments, the compound of Formula (A) is administered at adosage of about 40 mg/day, about 140 mg/day, about 280 mg/day about 420mg/day, about 560 mg/day, or about 840 mg/day. In some embodiments, thecompound of Formula (A) is administered in combination with additionaltherapeutic agents. In some embodiments, the additional therapeuticagent is a corticosteroid. In some embodiments, the additionaltherapeutic agent is cyclosporine (CSA), mycophenolate mofetil (MMF) ora combination thereof. In some embodiments, the compound of Formula (A)is administered orally. In some embodiments, the compound of Formula (A)is administered from day 1 to about day 120 following allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, thecompound of Formula (A) is administered from day 1 to about day 1000following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the patient is administered one or more donorlymphocyte infusions (DLI). In some embodiments, the DLI comprises CD3+lymphocytes. In some embodiments, the patient is administered one ormore donor lymphocyte infusions (DLI) following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, thecompound of Formula (A) is administered concurrently with a DLIfollowing allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the compound of Formula (A) is administered priorto a DLI following an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the compound of Formula (A) isadministered following a DLI following an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the compound ofFormula (A) is ibrutinib.

In some embodiments, there are provided uses of a compound of Formula(A) for preventing the occurrence of graft versus host disease (GVHD) orreducing the severity of GVHD occurrence in a patient requiring celltransplantation, wherein Formula (A) has the structure:

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—, —S(═O)₂NH—,—NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—,—NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-, aryl-,—NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or—C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or

two R₁₀ groups can together form a 5-, 6-, 7-, or 8-memberedheterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof. Insome embodiments, L₃, X and L₄ taken together form a nitrogen containingheterocyclic ring. In some embodiments, the nitrogen containingheterocyclic ring is a piperidine group. In some embodiments, G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematological malignancy. In some embodiments, the patienthas a relapsed or refractory hematological malignancy. In someembodiments, the patient has a B-cell malignancy. In some embodiments,the patient has a T-cell malignancy. In some embodiments, the patienthas a leukemia, a lymphoma, or a myeloma. In some embodiments, theB-cell malignancy is a non-Hodgkin's lymphoma. In some embodiments, theB-cell malignancy is chronic lymphocytic leukemia (CLL). In someembodiments, the B-cell malignancy is a relapsed or refractory B-cellmalignancy. In some embodiments, the B-cell malignancy is a relapsed orrefractory non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is a relapsed or refractory CLL. In some embodiments, thepatient has high risk CLL. In some embodiments, the patient has a 17pchromosomal deletion. In some embodiments, the patient has 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLL as determined by bonemarrow biopsy. In some embodiments, the patient has received one or moreprior anticancer agents. In some embodiments, the anticancer agent isselected from among alemtuzumab, bendamustine, bortezomib, CAL-101,chlorambucil, cyclophosphamide, dexamethasone, docetaxel, doxorubicin,endostatineverolimus, etoposide, fludarabine, fostamatinib,hydroxydaunorubicin, ibritumomab, ifosphamide, lenalidomide, mesalazine,ofatumumab, paclitaxel, pentostatin, prednisone, rituximab,temsirolimus, thalidomide, tositumomab, vincristine, or a combinationthereof. In some embodiments, the anticancer agent is rituximab. In someembodiments, the anticancer agent is alemtuzumab. In some embodiments,the anticancer agent is fludarabine, cyclophosphamide, and rituximab(FCR). In some embodiments, the anticancer agent is oxaliplatin,fludarabine, cytarabine, rituximab (OFAR). In some embodiments, theamount of the ACK inhibitor compound (e.g., a compound of Formula (A))prevents or reduces GVHD while maintaining a graft-versus-leukemia (GVL)reaction effective to reduce or eliminate the number of cancerous cellsin the blood of the patient. In some embodiments, the celltransplantation is a hematopoietic cell transplantation. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Insome embodiments, the GVHD is steroid resistant GVHD. In someembodiments, the GVHD is cyclosporin-resistant GVHD. In someembodiments, the GVHD is refractory GVHD. In some embodiments, the GHVDis oral GVHD. In some embodiments, the oral GVHD is reticular oral GVHD.In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered concurrently with an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered subsequent to an allogeneicbone marrow or hematopoietic stem cell transplant. In some embodiments,the patient is a candidate for receiving HLA-mismatched hematopoieticstem cells. In some embodiments, the patient is a candidate forreceiving unrelated donor hematopoietic stem cells, umbilical veinhematopoietic stem cells, or peripheral blood stem cells. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered orally. In some embodiments, the ACK inhibitorcompound (e.g., a compound of Formula (A)) is administered at a dosageof between about 0.1 mg/kg per day to about 100 mg/kg per day. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered at a dosage of about 40 mg/day, about 140 mg/day,about 280 mg/day, about 420 mg/day, about 560 mg/day, or about 840mg/day. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered in combination with otherprophylactic agents. In some embodiments, the ACK inhibitor compound(e.g., a compound of Formula (A)) is administered from day 1 to aboutday 120 following allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered from day 1 to about day 1000following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered in combination with one or more additionaltherapeutic agents. In some embodiments, the additional therapeuticagent is a corticosteroid. In some embodiments, the therapeutic agent iscyclosporine (CSA), mycophenolate mofetil (MMF) or a combinationthereof. In some embodiments, the patient has or will receive a donorlymphocyte infusions (DLI),In some embodiments, the patient isadministered one or more DLIs. In some embodiments, the patient isadministered two or more DLIs. In some embodiments, the DLI comprisesCD3+ lymphocytes. In some embodiments, the patient is administered oneor more donor lymphocyte infusions (DLI) following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, theACK inhibitor compound (e.g., a compound of Formula (A)) is administeredconcurrently with a DLI following allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to a DLI following an allogeneic bone marrow or hematopoietic stemcell transplant. In some embodiments, the ACK inhibitor compound (e.g.,a compound of Formula (A)) is administered following a DLI following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is ibrutinib.

In some embodiments, there are provided uses of a compound of Formula(A) with allogeneic hematopoietic stem cells and/or allogeneic T-cellsfor treating a patient for alleviation of a bone marrow mediateddisease, with alleviation of consequently developed graft versus hostdisease (GVHD), wherein the compound of Formula (A) has the structure:

wherein:

A is N;

R₁ is phenyl-O-phenyl or phenyl-S-phenyl;

R₂ and R₃ are independently H;

R₄ is L₃-X-L₄-G, wherein,

L₃ is optional, and when present is a bond, optionally substituted orunsubstituted alkyl, optionally substituted or unsubstituted cycloalkyl,optionally substituted or unsubstituted alkenyl, optionally substitutedor unsubstituted alkynyl;

X is optional, and when present is a bond, —O—, —C(═O)—, —S—, —S(═O)—,—S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—, —C(O)NR₉—, —S(═O)₂NH—,—NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—,—NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl-, aryl-,—NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or—C(═NR₁₁)O—;

L₄ is optional, and when present is a bond, substituted or unsubstitutedalkyl, substituted or unsubstituted cycloalkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted aryl, substituted or unsubstituted heteroaryl,substituted or unsubstituted heterocycle;

or L₃, X and L₄ taken together form a nitrogen containing heterocyclicring;

G is

wherein,

R₆, R₇ and R₈ are independently selected from among H, halogen, CN, OH,substituted or unsubstituted alkyl or substituted or unsubstitutedheteroalkyl or substituted or unsubstituted cycloalkyl, substituted orunsubstituted heterocycloalkyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl;

each R₉ is independently selected from among H, substituted orunsubstituted lower alkyl, and substituted or unsubstituted lowercycloalkyl;

each R₁₀ is independently H, substituted or unsubstituted lower alkyl,or substituted or unsubstituted lower cycloalkyl; or

two R₁₀ groups can together form a 5-, 6-, 7-, or 8-memberedheterocyclic ring; or

R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclicring; or

each R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof, andis administered prior to, concurrently with, or following theadministration of the allogeneic hematopoietic stem cells and/orallogeneic T-cells. In some embodiments, L₃, X and L₄ taken togetherform a nitrogen containing heterocyclic ring. In some embodiments, thenitrogen containing heterocyclic ring is a piperidine group. In someembodiments, G is

In some embodiments, the compound of Formula (A) is1-[(3R)-3[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one.In some embodiments, the patient has cancer. In some embodiments, thepatient has a hematological malignancy. In some embodiments, the patienthas a relapsed or refractory hematological malignancy. In someembodiments, the patient has a B-cell malignancy. In some embodiments,the patient has a T-cell malignancy. In some embodiments, the patienthas a leukemia, a lymphoma, or a myeloma. In some embodiments, theB-cell malignancy is a non-Hodgkin's lymphoma. In some embodiments, theB-cell malignancy is chronic lymphocytic leukemia (CLL). In someembodiments, the B-cell malignancy is a relapsed or refractory B-cellmalignancy. In some embodiments, the B-cell malignancy is a relapsed orrefractory non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is a relapsed or refractory CLL. In some embodiments, thepatient has high risk CLL. In some embodiments, the patient has a 17pchromosomal deletion. In some embodiments, the patient has 10%, 20%,30%, 40%, 50%, 60%, 70%, 80%, 90%, or greater CLL as determined by bonemarrow biopsy. In some embodiments, the patient has received one or moreprior anticancer agents. In some embodiments, the anticancer agent isselected from among alemtuzumab, bendamustine, bortezomib, CAL-101,chlorambucil, cyclophosphamide, dexamethasone, docetaxel, doxorubicin,endostatineverolimus, etoposide, fludarabine, fostamatinib,hydroxydaunorubicin, ibritumomab, ifosphamide, lenalidomide, mesalazine,ofatumumab, paclitaxel, pentostatin, prednisone, rituximab,temsirolimus, thalidomide, tositumomab, vincristine, or a combinationthereof. In some embodiments, the anticancer agent is rituximab. In someembodiments, the anticancer agent is alemtuzumab. In some embodiments,the anticancer agent is fludarabine, cyclophosphamide, and rituximab(FCR). In some embodiments, the anticancer agent is oxaliplatin,fludarabine, cytarabine, rituximab (OFAR). In some embodiments, theamount of the ACK inhibitor compound (e.g., a compound of Formula (A))prevents or reduces GVHD while maintaining a graft-versus-leukemia (GVL)reaction effective to reduce or eliminate the number of cancerous cellsin the blood of the patient. In some embodiments, the celltransplantation is a hematopoietic cell transplantation. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Insome embodiments, the GVHD is steroid resistant GVHD. In someembodiments, the GVHD is cyclosporin-resistant GVHD. In someembodiments, the GVHD is refractory GVHD. In some embodiments, the GHVDis oral GVHD. In some embodiments, the oral GVHD is reticular oral GVHD.In some embodiments, the oral GVHD is erosive oral GVHD. In someembodiments, the oral GVHD is ulcerative oral GVHD. In some embodiments,the oral GVHD is GVHD of the oral cavity. In some embodiments, the oralGVHD is GVHD of the oropharyngeal region. In some embodiments, the oralGVHD is GVHD of the pharyngeal region. In some embodiments, the oralGVHD is GVHD of the esophageal region. In some embodiments, the oralGVHD is acute oral GVHD. In some embodiments, the oral GVHD is chronicoral GVHD. In some embodiments, the patient exhibits one or moresymptoms of GVHD. In some embodiments, the patient has or will receivean allogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered concurrently with an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered subsequent to an allogeneicbone marrow or hematopoietic stem cell transplant. In some embodiments,the patient is a candidate for receiving HLA-mismatched hematopoieticstem cells. In some embodiments, the patient is a candidate forreceiving unrelated donor hematopoietic stem cells, umbilical veinhematopoietic stem cells, or peripheral blood stem cells. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered orally. In some embodiments, the ACK inhibitorcompound (e.g., a compound of Formula (A)) is administered at a dosageof between about 0.1 mg/kg per day to about 100 mg/kg per day. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is administered at a dosage of about 40 mg/day, about 140 mg/day,about 280 mg/day, about 420 mg/day, about 560 mg/day, or about 840mg/day. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered in combination with otherprophylactic agents. In some embodiments, the ACK inhibitor compound(e.g., a compound of Formula (A)) is administered from day 1 to aboutday 120 following allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the ACK inhibitor compound (e.g., acompound of Formula (A)) is administered from day 1 to about day 1000following allogeneic bone marrow or hematopoietic stem cell transplant.In some embodiments, the ACK inhibitor compound (e.g., a compound ofFormula (A)) is administered in combination with one or more additionaltherapeutic agents. In some embodiments, the additional therapeuticagent is a corticosteroid. In some embodiments, the therapeutic agent iscyclosporine (CSA), mycophenolate mofetil (MMF) or a combinationthereof. In some embodiments, the patient has or will receive a donorlymphocyte infusions (DLI),In some embodiments, the patient isadministered one or more DLIs. In some embodiments, the patient isadministered two or more DLIs. In some embodiments, the DLI comprisesCD3+ lymphocytes. In some embodiments, the patient is administered oneor more donor lymphocyte infusions (DLI) following an allogeneic bonemarrow or hematopoietic stem cell transplant. In some embodiments, theACK inhibitor compound (e.g., a compound of Formula (A)) is administeredconcurrently with a DLI following allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, the ACKinhibitor compound (e.g., a compound of Formula (A)) is administeredprior to a DLI following an allogeneic bone marrow or hematopoietic stemcell transplant. In some embodiments, the ACK inhibitor compound (e.g.,a compound of Formula (A)) is administered following a DLI following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the ACK inhibitor compound (e.g., a compound of Formula(A)) is ibrutinib.

Certain Terminology

It is to be understood that the foregoing general description and thefollowing detailed description are exemplary and explanatory only andare not restrictive of any subject matter claimed. In this application,the use of the singular includes the plural unless specifically statedotherwise. It must be noted that, as used in the specification and theappended claims, the singular forms “a,” “an” and “the” include pluralreferents unless the context clearly dictates otherwise. In thisapplication, the use of “or” means “and/or” unless stated otherwise.Furthermore, use of the term “including” as well as other forms, such as“include”, “includes,” and “included,” is not limiting.

As used herein, “ACK” and “Accessible Cysteine Kinase” are synonyms.They mean a kinase with an accessible cysteine residue. ACKs include,but are not limited to, BTK, ITK, Bmx/ETK, TEC, EFGR, HER4, HER4, LCK,BLK, C-src, FGR, Fyn, HCK, Lyn, YES, ABL, Brk, CSK, FER, JAK3, SYK. Insome embodiments, the ACK is a TEC family kinase. In some embodiments,the ACK is HER4. In some embodiments, the ACK is BTK. In someembodiments, the ACK is ITK.

As used herein, “amelioration” refers to any lessening of severity,delay in onset, slowing of growth, slowing of metastasis, or shorteningof duration of HER2-amplified breast cancer, whether permanent ortemporary, lasting or transient that can be attributed to or associatedwith administration of the compound or composition.

The term “Bruton's tyrosine kinase,” as used herein, refers to Bruton'styrosine kinase from Homo sapiens, as disclosed in, e.g., U.S. Pat. No.6,326,469 (GenBank Accession No. NP_000052).

The term “Bruton's tyrosine kinase homolog,” as used herein, refers toorthologs of Bruton's tyrosine kinase, e.g., the orthologs from mouse(GenBank Accession No. AAB47246), dog (GenBank Accession No. XP 549139),rat (GenBank Accession No. NP_001007799), chicken (GenBank Accession No.NP_989564), or zebra fish (GenBank Accession No. XP 698117), and fusionproteins of any of the foregoing that exhibit kinase activity towardsone or more substrates of Bruton's tyrosine kinase (e.g., a peptidesubstrate having the amino acid sequence “AVLESEEELYSSARQ” SEQ ID NO:1).

The term “HER4”, also known as ERBB4, also known as “V-erb-aerythroblastic leukemia viral oncogene homolog 4” means either (a) thenucleic acid sequence encoding a receptor tyrosine kinase that is amember of the epidermal growth factor receptor subfamily, or (b) theprotein thereof. For the nucleic acid sequence that comprises the humanHER4 gene see GenBank Accession No. NM_001042599. For the amino acidsequence that comprises the human HER4 protein see GenBank Accession No.NP_001036064.

The term “homologous cysteine,” as used herein refers to a cysteineresidue found within a sequence position that is homologous to that ofcysteine 481 of Bruton's tyrosine kinase, as defined herein. Forexample, cysteine 482 is the homologous cysteine of the rat ortholog ofBruton's tyrosine kinase; cysteine 479 is the homologous cysteine of thechicken ortholog; and cysteine 481 is the homologous cysteine in thezebra fish ortholog. In another example, the homologous cysteine of TXK,a Tec kinase family member related to Bruton's tyrosine, is Cys 350.

The term “irreversible BTK inhibitor,” as used herein, refers to aninhibitor of BTK that can form a covalent bond with an amino acidresidue of BTK. In one embodiment, the irreversible inhibitor of BTK canform a covalent bond with a Cys residue of BTK; in particularembodiments, the irreversible inhibitor can form a covalent bond with aCys 481 residue (or a homolog thereof) of BTK or a cysteine residue inthe homologous corresponding position of another tyrosine kinase, asshown in FIG. 7.

As used herein, the term “pERK” refers to phosphorylated ERK1 and ERK2at Thr202/Tyr 204 as detected by commercially available phospho-specificantibodies (e.g. Cell Signaling Technologies #4377).

The terms “individual”, “patient” and “subject” are usedinterchangeably. These terms refer to a mammal (e.g., a human) which isthe object of treatment, or observation. The term is not to be construedas requiring the supervision of a medical practitioner (e.g., aphysician, physician's assistant, nurse, orderly, or hospice careworker).

The terms “treat,” “treating” or “treatment”, as used herein, includelessening of severity of GVHD, delay in onset of GVHD, causingregression of GVHD, relieving a condition caused by of GVHD, or stoppingsymptoms which result from GVHD. The terms “treat,” “treating” or“treatment”, include, but are not limited to, prophylactic and/ortherapeutic treatments.

As used herein, oral GVHD refers to local manifestation of GVHD in theoral cavity, oropharynx, pharyngeal, or esophageal regions.

Graft Versus Host Disease

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administering tothe patient a composition comprising a therapeutically-effective amountof an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as,ibrutinib). In some embodiments, the patient requires hematopoietic celltransplantation. In some embodiments, the patient requires peripheralblood stem cell transplantation. In some embodiments, the patientrequires bone marrow transplantation. In some embodiments, the ACKinhibitor compound is administered prior to administration of the celltransplant. In some embodiments, the ACK inhibitor compound isadministered subsequent to administration of the cell transplant. Insome embodiments, the ACK inhibitor compound is administeredconcurrently with administration of the cell transplant. In someembodiments, the patient exhibits one or more symptoms of GVHD. In someembodiments, the patient exhibits one or more symptoms of acute GVHD. Insome embodiments, the patient exhibits one or more symptoms of chronicGVHD. In some embodiments, the GVHD is sclerodermatous GVHD. Exemplarysymptoms of GVHD include, but are not limited to, skin rash or reddenedareas on the skin, raised skin, blistering, thickening or tightening ofthe skin, yellow discoloration of the skin and/or eyes, abnormal bloodtest results, nausea, vomiting, diarrhea, abdominal swelling, abdominalcramping, increased dryness or irritation of the eyes, vision changes,dry mouth, white patches inside the mouth, pain or sensitivity to spicyfoods, shortness of breath, difficulty swallowing, pain with swallowing,weight loss, fatigue, muscle weakness, muscle pain, increased urinaryfrequency, burning or bleeding with urination, vaginal dryness ortightening, or penile dysfunction.

In some embodiments, the patient exhibits one or more symptoms of oralGVHD. In some embodiments, the patient exhibits one or more symptoms ofacute oral GVHD. In some embodiments, the patient exhibits one or moresymptoms of chronic oral GVHD. Exemplary symptoms of oral GVHD include,but are not limited to, oral tissue inflammation, dry mouth, punctate orgeneralized mucosal erythema, white striae or papules on the oral mucosaand lips, mucosal erosion-desquamation-ulceration, pain or sensitivityto spicy foods, difficulty swallowing, pain with swallowing,pharyngo-esophageal stricture, xerostomia, lichen planus, poor boluscontrol, pharyngeal retention, excessive mucous secretion, oral tissueinflammation, and ulceration. In some embodiments, the patient suffersfrom refractory GVHD. In some embodiments, the oral GVHD is reticularoral GVHD. In some embodiments, the oral GVHD is erosive oral GVHD. Insome embodiments, the oral GVHD is ulcerative oral GVHD.

In some embodiments, the patient suffers from steroid resistant GVHD. Insome embodiments, the steroid resistant GVHD is acute GVHD. In someembodiments, the steroid resistant GVHD is chronic GVHD. In someembodiments, the patient suffers from cyclosporin-resistant GVHD.

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring stem cell transplantation comprisingadministering to the patient a composition comprising atherapeutically-effective amount of an ACK inhibitor compound (e.g., anITK or BTK inhibitor, such as for example ibrutinib). In someembodiments, the patient requires hematopoietic stem celltransplantation. In some embodiments, the patient requires peripheralblood stem cell transplantation. In some embodiments, the patientrequires bone marrow transplantation. In some embodiments, the ACKinhibitor compound is administered prior to administration of the stemcell transplant. In some embodiments, the ACK inhibitor compound isadministered subsequent to administration of the stem cell transplant.In some embodiments, the ACK inhibitor compound is administeredconcurrently with administration of the stem cell transplant. In someembodiments, the ACK inhibitor compound is administered prior to,subsequent to, or concurrently with administration of allogeneichematopoietic stem cells and/or allogeneic T-cells.

Further described herein are methods of treating a patient foralleviation of a bone marrow mediated disease, with alleviation ofconsequently developed graft versus host disease (GVHD), comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, wherein a therapeutically effective amount of an ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as, ibrutinib)is administered prior to, subsequently, or concurrently withadministration of the allogeneic hematopoietic stem cells and/orallogeneic T-cells.

Treatment of proliferative blood disorders, such as leukemia, lymphomaand myeloma usually involves one or more forms of chemotherapy and/orradiation therapy. These treatments destroy malignant cells, but alsodestroy healthy blood cells. Allogeneic hematopoietic celltransplantation is an effective therapy for the treatment of manyhematologic malignancies, including, for example, B-cell and T-cellmalignancies. In allogeneic hematopoietic cell transplantation, bonemarrow (or, in some cases, peripheral blood) from an unrelated or arelated (but not identical twin) donor is used to replace the healthyblood cells destroyed in the cancer patient. The bone marrow (orperipheral blood) contains stem cells, which are the precursors to allthe different cell types (e.g., red cells, phagocytes, platelets andlymphocytes) found in blood. Allogeneic hematopoietic celltransplantation is known to have both a restorative effect and acurative effect. The restorative effect arises from the ability of thestem cells to repopulate the cellular components of blood. The curativeproperties of allogeneic hematopoietic cell transplantation derivelargely from a graft-versus-leukemia (GVL) effect. The transplantedhematopoietic cells from the donor (specifically, the T lymphocytes)attack the cancerous cells, enhancing the suppressive effects of theother forms of treatment. Essentially, the GVL effect comprises anattack on the cancerous cells by the blood cells derived from thetransplantation, making it less likely that the malignancy will returnafter transplant. Controlling the GVL effect prevents escalation of theGVL effect into GVHD. A similar effect against tumors (graft-versustumor) is also known.

Allogeneic hematopoietic cell transplantation is often toxic to thepatient. This toxicity arises from the difficulty in dissociating theGVL or GVT effect from graft-versus-host disease (GVHD), an often-lethalcomplication of allogeneic BMT.

GVHD is a major complication of allogeneic hematopoietic cell transplant(HCT). GVHD is an inflammatory disease initiated by T cells in the donorgraft that recognize histocompatibility and other tissue antigens of thehost and GVHD is mediated by a variety of effector cells andinflammatory cytokines. GVHD presents in both acute and chronic forms.The most common symptomatic organs are the skin, liver, andgastrointestinal tract, including the oral cavity and oropharyngealregions. GVHD may involve other organs such as the lung. Treatment ofGVHD is generally only 50-75% successful; the remainder of patientsgenerally do not survive. The risk and severity of this immune-mediatedcondition are directly related to the degree of mismatch between a hostand the donor of hematopoietic cells. For example, GVHD develops in upto 30% of recipients of human leukocyte antigen (HLA)-matched siblingmarrow, in up to 60% of recipients of HLA-matched unrelated donormarrow, and in a higher percentage of recipient of HLA-mismatchedmarrow. Patients with mild intestinal GVHD present with anorexia,nausea, vomiting, abdominal pain and diarrhea, whereas patients withsevere GVHD are disabled by these symptoms. If untreated, symptoms ofintestinal GVHD persist and often progress; spontaneous remissions areunusual. In its most severe form, GVHD leads to necrosis and exfoliationof most of the epithelial cells of the intestinal mucosa, a frequentlyfatal condition. The symptoms of acute GVHD usually present within 100days of transplantation. The symptoms of chronic GVHD usually presentsomewhat later, up to three years after allogeneic HCT, and are oftenproceeded by a history of acute GVHD.

Oral manifestations of GVHD are seen in both acute GVHD (aGVHD) andchronic GVHD (cGVHD). Oral involvement ranges between 33% and 75% forpatients with aGVHD and up to about 80% for those with cGVHD.Involvement of the salivary glands may cause dryness of the oral mucosaand oral pain may be the first presenting symptom. Oral lesions in GVHDmay be lichenoid or lupus-like in appearance. Oral findings of aGVHDinclude painful desquamative, erythematous, and ulcerative mucosallesions. In cGVHD, they are lichenoid with associated erythema andulcerations; additionally, they may be associated with sicca syndromecharacterized by xerostomia and progressive salivary gland atrophy. Oralcomplications include pain due to the mucosal changes, altered orreduced taste, and may have a potential impact on speech, deglutition,and use of oral prostheses (when present). Oral infection, particularlydue to Candida species, and dental demineralization and caries may alsooccur. Oral manifestations of cGVHD can significantly affect the lifequality of patients through discomfort and impairment of the oral intakeleading to malnutrition and increased morbidity.

The conventional management of oral cGVHD consists of systemicimmunosuppressive therapies combined with proper oral hygiene and thejudicious use of topical steroids. However, for patients with oral cGVHDas the most significant clinical finding, the use of systemicimmunosuppressants may result in immunosuppression of the host withattendant systemic complications. In addition, some patients experienceconsiderable and refractory oral complications, even with maximum dosesof systemic immunosuppressants.

First-line therapy of oral GVHD is mostly systemic in nature, consistingof cyclosporin and steroids. The most common salvage treatments forcGVHD are thalidomide, tacrolimus, mycophenolate mofetil, T celldepletion by Campath-1, and phototherapy. Oral GVHD is often refractoryto conventional treatment and therefore complementary topical treatmentis required. Several agents are currently used for local treatments suchas palliative rinses, topical immunosuppressive agents, thalidomide,retinoids, and phototherapy for oral GVHD.

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administering tothe patient a composition comprising a therapeutically-effective amountof an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib). In some embodiments, the patient requireshematopoietic cell transplantation. Further described herein are methodsof treating a patient for alleviation of a bone marrow mediated disease,with alleviation of consequently developed graft versus host disease(GVHD), comprising administering to the patient allogeneic hematopoieticstem cells and/or allogeneic T-cells, wherein a therapeuticallyeffective amount of an ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as ibrutinib) is administered prior to, concurrentlywith, or following the allogeneic hematopoietic stem cells and/orallogeneic T-cells. In some embodiments, the patient has cancer. In someembodiments, the patient has a hematologic malignancy. In someembodiments, the patient has a B-cell malignancy. In some embodiments,the patient has a T-cell malignancy. In some embodiments, the patienthas a leukemia, lymphoma, or a myeloma. In some embodiments, the B-cellmalignancy is a non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is chronic lymphocytic leukemia (CLL). In some embodiments,the B-cell malignancy is a relapsed or refractory B-cell malignancy. Insome embodiments, the B-cell malignancy is a relapsed or refractorynon-Hodgkin's lymphoma. In some embodiments, the B-cell malignancy is arelapsed or refractory CLL. In some embodiments, the patient has highrisk CLL. In some embodiments, the patient has a 17p chromosomaldeletion. In some embodiments, the patient has 10%, 20%, 30%, 40%, 50%,60%, 70%, 80%, 90%, or greater CLL as determined by bone marrow biopsy.In some embodiments, the patient has received one or more prioranticancer agents. In some embodiments, the anticancer agent is selectedfrom among alemtuzumab, bendamustine, bortezomib, CAL-101, chlorambucil,cyclophosphamide, dexamethasone, docetaxel, doxorubicin,endostatineverolimus, etoposide, fludarabine, fostamatinib,hydroxydaunorubicin, ibritumomab, ifosphamide, lenalidomide, mesalazine,ofatumumab, paclitaxel, pentostatin, prednisone, rituximab,temsirolimus, thalidomide, tositumomab, vincristine, or a combinationthereof. In some embodiments, the anticancer agent is rituximab. In someembodiments, the anticancer agent is alemtuzumab. In some embodiments,the anticancer agent is fludarabine, cyclophosphamide, and rituximab(FCR). In some embodiments, the anticancer agent is oxaliplatin,fludarabine, cytarabine, rituximab (OFAR). In some embodiments, acompound disclosed herein prevents or reduces GVHD while maintaining agraft-versus-leukemia (GVL) reaction effective to reduce or eliminatethe number of cancerous cells in the blood of the patient. In someembodiments, the GVHD is acute GVHD. In some embodiments, the GVHD ischronic GVHD. In some embodiments, the GHVD is oral GVHD. In someembodiments, the oral GVHD is GVHD of the oral cavity. In someembodiments, the oral GVHD is GVHD of the oropharyngal region. In someembodiments, the oral GVHD is GVHD of the pharyngeal region. In someembodiments, the oral GVHD is GVHD of the esophageal region. In someembodiments, the oral GVHD is acute oral GVHD. In some embodiments, theoral GVHD is chronic oral GVHD. In some embodiments, the patient has orwill receive an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, an ACK inhibitor compound disclosedherein is administered concurrently with an allogeneic bone marrow orhematopoietic stem cell transplant. In some embodiments, an ACKinhibitor compound disclosed herein is administered prior to anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, an ACK inhibitor compound disclosed herein is administeredsubsequent to an allogeneic bone marrow or hematopoietic stem celltransplant. In some embodiments, the patient is a candidate forreceiving HLA-mismatched hematopoietic stem cells. In some embodiments,the patient is a candidate for receiving unrelated donor hematopoieticstem cells, umbilical vein hematopoietic stem cells, or peripheral bloodstem cells. In some embodiments, an ACK inhibitor compound disclosedherein is administered subsequent to a patient exhibiting one or moresymptoms of oral GVHD, wherein the patient an allogeneic bone marrow orhematopoietic stem cell transplant.

In some embodiments, the patient is administered a donor lymphocyteinfusions (DLI). A donor lymphocyte infusion is a blood cell infusion inwhich CD3+ lymphocytes from the original stem cell donor are infused,after the transplant, to augment an anti-tumor immune response or ensurethat the donor stem cells remain engrafted. These donated white bloodcells contain cells of the immune system that can recognize and destroycancer cells. In some embodiments, the therapy induces a remission ofthe patient's cancer by a graft-versus-tumor effect (GVT). In someembodiments, the donor T-cells can attack and control the growth ofresidual cancer cells providing the GVT effect. In some embodiments, thepatient is administered one or more donor lymphocyte infusions (DLI). Insome embodiments, the DLI comprises CD3+ lymphocytes. In someembodiments, the patient is administered one or more donor lymphocyteinfusions (DLI) following an allogeneic bone marrow or hematopoieticstem cell transplant. In some embodiments, the compound of Formula (A)is administered concurrently with a DLI following allogeneic bone marrowor hematopoietic stem cell transplant. In some embodiments, the compoundof Formula (A) is administered prior to a DLI following an allogeneicbone marrow or hematopoietic stem cell transplant. In some embodiments,the compound of Formula (A) is administered following a DLI following anallogeneic bone marrow or hematopoietic stem cell transplant. In someembodiments, the compound of Formula (A) is ibrutinib.

In some embodiments, the patient has a non-Hodgkin's lymphoma. In someembodiments, the patient has a Hodgkin's lymphoma. In some embodiments,the patient has a B-cell malignancy. In some embodiments, the B-cellmalignancy is chronic lymphocytic leukemia (CLL), small lymphocyticlymphoma (SLL), diffuse large B-cell lymphoma (DLBCL), follicularlymphoma (FL), activated B-cell diffuse large B-cell lymphoma(ABC-DLBCL), germinal center diffuse large B-cell lymphoma (GCB DLBCL),primary mediastinal B-cell lymphoma (PMBL), Burkitt's lymphoma,immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma,mantle cell lymphoma (MCL), B cell prolymphocytic leukemia,lymphoplasmacytic lymphoma, Waldenström macroglobulinemia, splenicmarginal zone lymphoma, plasma cell myeloma, plasmacytoma, extranodalmarginal zone B cell lymphoma, nodal marginal zone B cell lymphoma,mediastinal (thymic) large B cell lymphoma, intravascular large B celllymphoma, primary effusion lymphoma, or lymphomatoid granulomatosis. Insome embodiments, the patient has a T-cell malignancy. In someembodiments, the T-cell malignancy is peripheral T-cell lymphoma nototherwise specified (PTCL-NOS), anaplastic large cell lymphoma,angioimmunoblastic lymphoma, cutaneous T-cell lymphoma, adult T-cellleukemia/lymphoma (ATLL), blastic NK-cell lymphoma, enteropathy-typeT-cell lymphoma, hematosplenic gamma-delta T-cell lymphoma,lymphoblastic lymphoma, nasal NK/T-cell lymphomas, or treatment-relatedT-cell lymphomas. In some embodiments, the subject has multiple myeloma.

In some embodiments, the patient has a relapsed or refractoryhematologic cancer. In some embodiments, the relapsed or refractoryhematologic cancer is a leukemia, a lymphoma, or a myeloma. In someembodiments, the relapsed or refractory hematologic cancer is anon-Hodgkin's lymphoma. In some embodiments, the relapsed or refractoryhematologic cancer is a Hodgkin's lymphoma. In some embodiments, therelapsed or refractory hematologic cancer is a B-cell malignancy. Insome embodiments, the B-cell malignancy is chronic lymphocytic leukemia(CLL), small lymphocytic lymphoma (SLL), diffuse large B-cell lymphoma(DLBCL), follicular lymphoma (FL), activated B-cell diffuse large B-celllymphoma (ABC-DLBCL), germinal center diffuse large B-cell lymphoma (GCBDLBCL), primary mediastinal B-cell lymphoma (PMBL), Burkitt's lymphoma,immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma,mantle cell lymphoma (MCL), B cell prolymphocytic leukemia,lymphoplasmacytic lymphoma, Waldenström macroglobulinemia, splenicmarginal zone lymphoma, plasma cell myeloma, plasmacytoma, extranodalmarginal zone B cell lymphoma, nodal marginal zone B cell lymphoma,mediastinal (thymic) large B cell lymphoma, intravascular large B celllymphoma, primary effusion lymphoma, or lymphomatoid granulomatosis. Insome embodiments, the relapsed or refractory hematologic cancer is aT-cell malignancy. In some embodiments, the T-cell malignancy isperipheral T-cell lymphoma not otherwise specified (PTCL-NOS),anaplastic large cell lymphoma, angioimmunoblastic lymphoma, cutaneousT-cell lymphoma, adult T-cell leukemia/lymphoma (ATLL), blastic NK-celllymphoma, enteropathy-type T-cell lymphoma, hematosplenic gamma-deltaT-cell lymphoma, lymphoblastic lymphoma, nasal NK/T-cell lymphomas, ortreatment-related T-cell lymphomas. In some embodiments, the subject hasa relapsed or refractory multiple myeloma. In some embodiments, thepatient has a B-cell malignancy. In some embodiments, the B-cellmalignancy is a non-Hodgkin's lymphoma. In some embodiments, the B-cellmalignancy is chronic lymphocytic leukemia (CLL). In some embodiments,the B-cell malignancy is a relapsed or refractory B-cell malignancy. Insome embodiments, the B-cell malignancy is a relapsed or refractorynon-Hodgkin's lymphoma. In some embodiments, the B-cell malignancy is arelapsed or refractory CLL.

In some embodiments, the patient exhibits one or more symptoms of ahematologic cancer. In some embodiments, the subject exhibits one ormore symptoms of a B-cell malignancy. In some embodiments, the subjectexhibits one or more symptoms of a leukemia, a lymphoma, or a myeloma.In some embodiments, the subject exhibits one or more symptoms such as,but not limited to, abnormal B-cell function, abnormal B-cell size orshape, abnormal B-cell count, fatigue, fever, night sweats, frequentinfection, enlarged lymph nodes, paleness, anemia, easy bleeding orbruising, loss of appetite, weight loss, bone or joint pain, headaches,and petechiae.

In some embodiments, the subject has a high risk of cancer recurrence.In some embodiments, the subject is a mammal, such as, but not limitedto a human, a non-human primate, mouse, rat, rabbit, goat, dog, cat, orcow. In some embodiments, the mammal is a human. In some embodiments, ahigh risk of cancer recurrence is determined based on the expression orpresence of a biomarker. In some embodiments, the biomarker includesPMSB1 P11A G/C heterozygote, CD68, suppressor of cytokine signaling 1(SOCS1), LIM domain only 2 (LMO2), CD137, or a combination thereof.

Combination Therapies

Described herein methods of preventing the occurrence of graft versushost disease (GVHD) or reducing the severity of GVHD occurrence in apatient requiring cell transplantation comprising co-administering tothe individual a composition comprising a therapeutically-effectiveamount of an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, suchas for example ibrutinib) and an additional therapeutic agent. Furtherdescribed herein are methods of treating a patient for alleviation of abone marrow mediated disease, with alleviation of consequently developedoral graft versus host disease (GVHD), comprising co-administering tothe individual a composition comprising a therapeutically-effectiveamount of an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, suchas for example ibrutinib) and an additional therapeutic agent prior to,subsequent to or concurrently with the allogeneic hematopoietic stemcells and/or allogeneic T-cells. In some embodiments, the individual isadministered an additional therapy such as, but not limited to,extracorporeal photopheresis or infusion of mesenchymal stem cells ordonor lymphocytes.

In some embodiments, the additional therapeutic agent is an anti-GVHDtherapeutic agent. In some embodiments, the anti-GVHD therapeutic agentis an immunosuppressive drug. In some embodiments, the immunosuppressivedrug includes cyclosporine, tacrolimus, methotrexate, mycophenolatemofetil, corticosteroids, azathioprine or antithymocyte globulin (ATG).In some embodiments, the immunosuppressive drug is a monoclonal antibody(for example, anti-CD3, anti-CD5, and anti-IL-2 antibodies). In someembodiments, the immunosuppressive drug is Mycophenolate mofetil,Alemtuzumab, Antithymocyte globulin (ATG), Sirolimus, Tacrolimus,Thalidomide, Daclizumab, Infliximab, or Clofazimine are of use to treatchronic GVHD. In some embodiments, the additional therapeutic agent isdenileukin diftitox, defibrotide, budesonide, beclomethasonedipropionate, or pentostatin.

In some embodiments, the additional therapeutic agent is an IL-6receptor inhibitor. In some embodiments, the additional therapeuticagent is an IL-6 receptor antibody.

In some embodiments, the additional therapeutic agent is a TLR5 agonist.

In some embodiments, the patient undergoes an additional therapy such asextracorporeal photopheresis or infusion of mesenchymal stem cells ordonor lymphocytes.

In some embodiments, the additional therapeutic agent is a topicallyactive corticosteroid (TAC). In some embodiments, the TAC isbeclomethasone dipropionate, alciometasone dipropionate, busedonide, 22Sbusesonide, 22R budesonide, beclomethasone-17-monopropionate,betamethasone, clobetasol propionate, dexamethasone, diflorasonediacetate, flunisolide, fluocinonide, flurandrenolide, fluticasonepropionate, halobetasol propionate, halcinocide, mometasone furoate,triamcinalone acetonide or a combination thereof.

In some embodiments, the additional therapeutic agent is an antifungalagent. In some embodiments, the additional therapeutic agent isnystatin, clotrimazole, amphotericin, fluconazole itraconazole or acombination thereof.

In some embodiments, the additional therapeutic agent is a sialogogue.In some embodiments, the additional therapeutic agent is cevimeline,pilocarpine, bethanechol or a combination thereof.

In some embodiments, the additional therapeutic agent is a topicalanesthetic. In some embodiments, the additional therapeutic agent islidocaine, dyclonine, diphenhydramine, doxepin or a combination thereof.

In the methods described herein, any suitable technique forchemotherapy, biotherapy, immunosuppression and radiotherapy known inthe art may be used. For example, the chemotherapeutic agent may be anyagent that exhibits an oncolytic effect against cancer cells orneoplastic cells of the subject. For example, the chemotherapeutic agentmay be, without limitation, an anthracycline, an alkylating agent, analkyl sulfonate, an aziridine, an ethylenimine, a methyhnelamine, anitrogen mustard, a nitrosourea, an antibiotic, an antimetabolite, afolic acid analogue, a purine analogue, a pyrimidine analogue, anenzyme, a podophyllotoxin, a platinum-containing agent or a cytokine.Preferably, the chemotherapeutic agent is one that is known to beeffective against the particular cell type that is cancerous orneoplastic. In some embodiments, the chemotherapeutic agent is effectivein the treatment of hematopoietic malignancies, such as thiotepa,cisplatin-based compounds, and cyclophosphamide. Cytokines includeinterferons, G-CSF, erythropoietin, GM-CSF, interleukins, parathyroidhormone, and the like. Biotherapies include alemtuzumab, rituximab,bevacizumab, vascular disrupting agents, lenalidomide, and the like.Radiosensitizers include nicotinomide, and the like.

In some embodiments, the ACK inhibitor is administered in combinationwith a chemotherapeutic agent or biologic agent selected from among anantibody, a B cell receptor pathway inhibitor, a T cell receptorinhibitor, a PI3K inhibitor, an IAP inhibitor, an mTOR inhibitor, aradioimmunotherapeutic, a DNA damaging agent, a histone deacetylaseinhibitor, a protein kinase inhibitor, a hedgehog inhibitor, an Hsp90inhibitor, a telomerase inhibitor, a Jak1/2 inhibitor, a proteaseinhibitor, an IRAK inhibitor, a PKC inhibitor, a PARP inhibitor, aCYP3A4 inhibitor, an AKT inhibitor, an Erk inhibitor, a proteosomeinhibitor, an alkylating agent, an anti-metabolite, a plant alkaloid, aterpenoid, a cytotoxin, a topoisomerase inhibitor, or a combinationthereof. In some embodiments, the B cell receptor pathway inhibitor is aCD79A inhibitor, a CD79B inhibitor, a CD19 inhibitor, a Lyn inhibitor, aSyk inhibitor, a PI3K inhibitor, a Blnk inhibitor, a PLCγ inhibitor, aPKCβ inhibitor, a CD22 inhibitor, a Bcl-2 inhibitor, an IRAK 1/4inhibitor, a JAK inhibitor (e.g., ruxolitinib, baricitinib, CYT387,lestauritinib, pacritinib, TG101348, SAR302503, tofacitinib (Xeljanz),etanercept (Enbrel), GLPG0634, R256), a microtubule inhibitor, a Topo IIinhibitor, anti-TWEAK antibody, anti-IL17 bispecific antibody, a CK2inhibitor, anaplastic lymphoma kinase (ALK) and c-Met inhibitors,demethylase enzyme inhibitors such as demethylase, HDM, LSDI and KDM,fatty acid synthase inhibitors such as spirocyclic piperidinederivatives, glucocorticosteriod receptor agonist, fusion anti-CD19-cytotoxic agent conjugate, antimetabolite, p70S6K inhibitor, immunemodulators, AKT/PKB inhibitor, procaspase-3 activator PAC-1, BRAFinhibitor, lactate dehydrogenase A (LDH-A) inhibitor, CCR2 inhibitor,CXCR4 inhibitor, chemokine receptor antagonists, DNA double strandedbreak repair inhibitors, NOR202, GA-101, TLR2 inhibitor, or acombination thereof. In some embodiments, the T cell receptor inhibitoris Muromonab-CD3. In some embodiments, the chemotherapeutic agent isselected from among rituximab (rituxan), carfilzomib, fludarabine,cyclophosphamide, vincristine, prednisalone. chlorambucil, ifosphamide,doxorubicin, mesalazine, thalidomide, revlimid, lenalidomide,temsirolimus, everolimus, fostamatinib, paclitaxel, docetaxel,ofatumumab, dexamethasone, bendamustine, prednisone, CAL-101,ibritumomab, tositumomab, bortezomib, pentostatin, endostatin,ritonavir, ketoconazole, an anti-VEGF antibody, herceptin, cetuximab,cisplatin, carboplatin, docetaxel, erlotinib, etopiside, 5-fluorouracil,gemcitabine, ifosphamide, imatinib mesylate (Gleevec), gefitinib,erlotinib, procarbazine, prednisone, irinotecan, leucovorin,mechlorethamine, methotrexate, oxaliplatin, paclitaxel, sorafenib,sunitinib, topotecan, vinblastine, GA-1101, dasatinib, Sipuleucel-T,disulfiram, epigallocatechin-3-gallate, salinosporamide A, ONX0912,CEP-18770, MLN9708, R-406, lenalinomide, spirocyclic piperidinederivatives, quinazoline carboxamide azetidine compounds, thiotepa,DWA2114R, NK121, IS 3 295, 254-S, alkyl sulfonates such as busulfan,improsulfan and piposulfan; aziridines such as benzodepa, carboquone,meturedepa and uredepa; ethylenimine, methylmelamines such asaltretamine, triethylenemelamine, triethylenephosphoramide,triethylenethiophosphoramide and trimethylmelamine; chlornaphazine;estramustine; ifosfamide; mechlorethamine; oxide hydrochloride;novobiocin; phenesterine; prednimustine; trofosfamide; uracil mustard;nitrosoureas such as carmustine, chlorozotocin, fotemustine, lomustine,nimustine, ranimustine; antibiotics such as aclacinomycins, actinomycin,anthramycin, azaserine, bleomycins, cactinomycin, calicheamicin,carubicin, carminomycin, carzinophilin, chromomycins, dactinomycin,daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin,epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins,mycophenolic acid, nogalamycin, olivomycins, peplomycin, porfiromycin,puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin,tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites such asmethotrexate and 5-fluorouracil (5-FU); folic acid analogues such asdenopterin, methotrexate, pteropterin, trimetrexate; purine analogs suchas fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidineanalogs such as ancitabine, azacitidine, 6-azauridine, carmofur,cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine;androgens such as calusterone, dromostanolone propionate, epitiostanol,mepitiostane, testolactone; anti-adrenals such as aminoglutethimide,mitotane, trilostane; folic acid replenisher such as folinic acid;aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine;bestrabucil; bisantrene; edatrexate; defosfamide; demecolcine;diaziquone; eflornithine; elliptinium acetate; etoglucid; galliumnitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone;mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllinicacid; 2-ethylhydrazide; procarbazine; polysaccharide-K; razoxane;sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2′,2″-trichlorotriethylamine; urethan; vindesine; dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; cytosinearabinoside; taxoids, e.g., paclitaxel and docetaxel; 6-thioguanine;mercaptopurine; methotrexate; platinum analogs; platinum; etoposide(VP-16); ifosfamide; mitomycin C; mitoxantrone; vincristine;vinorelbine; Navelbine; Novantrone; teniposide; daunomycin; aminopterin;Xeloda; ibandronate; CPT1 1; topoisomerase inhibitor RFS 2000;difluoromethylornithine (DMFO); retinoic acid; esperamycins;capecitabine; and pharmaceutically acceptable salts, acids orderivatives of; anti-hormonal agents such as anti-estrogens includingfor example tamoxifen, raloxifene, aromatase inhibiting 4(5)-imidazoles,4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone andtoremifene (Fareston); antiandrogens such as flutamide, nilutamide,bicalutamide, leuprolide and goserelin; ACK inhibitors such as AVL-263(Avila Therapeutics/Celgene Corporation), AVL-292 (AvilaTherapeutics/Celgene Corporation), AVL-291 (Avila Therapeutics/CelgeneCorporation), BMS-488516 (Bristol-Myers Squibb), BMS-509744(Bristol-Myers Squibb), CGI-1746 (CGI Pharma/Gilead Sciences), CTA-056,GDC-0834 (Genentech), HY-11066 (also, CTK4I7891, HMS3265G21, HMS3265G22,HMS3265H21, HMS3265H22, 439574-61-5, AG-F-54930), ONO-4059 (OnoPharmaceutical Co., Ltd.), ONO-WG37 (Ono Pharmaceutical Co., Ltd.),PLS-123 (Peking University), RN486 (Hoffmann-La Roche), HM71224 (HanmiPharmaceutical Company Limited) or a combination thereof.

When an additional agent is co-administered with an ACK inhibitor, theadditional agent and the ACK inhibitor do not have to be administered inthe same pharmaceutical composition, and are optionally, because ofdifferent physical and chemical characteristics, administered bydifferent routes. The initial administration is made, for example,according to established protocols, and then, based upon the observedeffects, the dosage, modes of administration and times of administrationare modified.

By way of example only, if a side effect experienced by an individualupon receiving an ACK inhibitor is nausea, then it is appropriate toadminister an anti-emetic agent in combination with the ACK inhibitor.

Or, by way of example only, the therapeutic effectiveness of an ACKinhibitor described herein is enhanced by administration of an adjuvant(i.e., by itself the adjuvant has minimal therapeutic benefit, but incombination with another therapeutic agent, the overall therapeuticbenefit to the patient is enhanced). Or, by way of example only, thebenefit experienced by an individual is increased by administering anACK inhibitor described herein with another therapeutic agent (whichalso includes a therapeutic regimen) that also has therapeutic benefit.In any case, regardless of the disease, disorder being treated, theoverall benefit experienced by the patient is in some embodiments simplyadditive of the two therapeutic agents or in other embodiments, thepatient experiences a synergistic benefit.

The particular choice of compounds used will depend upon the diagnosisof the attending physicians and their judgment of the condition of thepatient and the appropriate treatment protocol. The compounds areoptionally administered concurrently (e.g., simultaneously, essentiallysimultaneously or within the same treatment protocol) or sequentially,depending upon the nature of the disorder, the condition of the patient,and the actual choice of compounds used. The determination of the orderof administration, and the number of repetitions of administration ofeach therapeutic agent during a treatment protocol, is based on anevaluation of the disease being treated and the condition of thepatient.

In some embodiments, therapeutically-effective dosages vary when thedrugs are used in treatment combinations. Methods for experimentallydetermining therapeutically-effective dosages of drugs and other agentsfor use in combination treatment regimens are described in theliterature. For example, the use of metronomic dosing, i.e., providingmore frequent, lower doses in order to minimize toxic side effects, hasbeen described extensively in the literature Combination treatmentfurther includes periodic treatments that start and stop at varioustimes to assist with the clinical management of the patient.

For combination therapies described herein, dosages of theco-administered compounds will of course vary depending on the type ofco-drug employed, on the specific drug employed, on the disorder beingtreated and so forth. In addition, when co-administered with anadditional therapeutic agent, an ACK inhibitor described herein isadministered either simultaneously with the additional therapeuticagent, or sequentially. If administered sequentially, the attendingphysician will decide on the appropriate sequence of administeringprotein in combination with the biologically active agent(s).

If the additional therapeutic agent and the ACK inhibitor areadministered simultaneously, the multiple therapeutic agents areoptionally provided in a single, unified form, or in multiple forms (byway of example only, either as a single pill or as two separate pills).In some embodiments, one of the therapeutic agents is given in multipledoses, or both are given as multiple doses. If not simultaneous, thetiming between the multiple doses is from about more than zero weeks toless than about four weeks. In addition, the combination methods,compositions and formulations are not to be limited to the use of onlytwo agents; the use of multiple therapeutic combinations is alsoenvisioned.

It is understood that the dosage regimen to treat, prevent, orameliorate the condition(s) for which relief is sought, can be modifiedin accordance with a variety of factors. These factors include thedisorder from which the subject suffers, as well as the age, weight,sex, diet, and medical condition of the subject. Thus, the dosageregimen actually employed can vary widely and therefore can deviate fromthe dosage regimens set forth herein.

In some embodiments, the pharmaceutical agents which make up thecombination therapy disclosed herein are administered in a combineddosage form, or in separate dosage forms intended for substantiallysimultaneous administration. In some embodiments, the pharmaceuticalagents that make up the combination therapy are administeredsequentially, with either therapeutic compound being administered by aregimen calling for two-step administration. In some embodiments, thetwo-step administration regimen calls for sequential administration ofthe active agents or spaced-apart administration of the separate activeagents. The time period between the multiple administration steps rangesfrom a few minutes to several hours, depending upon the properties ofeach pharmaceutical agent, such as potency, solubility, bioavailability,plasma half-life and kinetic profile of the pharmaceutical agent. Insome embodiments, circadian variation of the target moleculeconcentration determines the optimal dose interval.

In some embodiments, the ACK inhibitor compound and the additionaltherapeutic agent are administered in a unified dosage form. In someembodiments, the ACK inhibitor compound and the additional therapeuticagent are administered in separate dosage forms. In some embodiments,the ACK inhibitor compound and the additional therapeutic agent areadministered simultaneously or sequentially.

Administration

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administering tothe patient a composition comprising a therapeutically-effective amountof an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib).

Further described herein are methods of reducing the severity of GVHDoccurrence in a patient requiring cell transplantation comprisingadministering to the patient a composition comprising atherapeutically-effective amount of an ACK inhibitor compound (e.g., anITK or BTK inhibitor, such as for example ibrutinib).

Further described herein are methods of treating a patient foralleviation of a bone marrow mediated disease, with alleviation ofconsequently developed graft versus host disease (GVHD), comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, wherein a therapeutically effective amount of an ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as for exampleibrutinib) is administered prior to or concurrently with the allogeneichematopoietic stem cells and/or allogeneic T-cells. In some embodiments,the ACK inhibitor compound is(R)-1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(i.e., PCI-32765/ibrutinib).

The ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib) is administered before, during or after thedevelopment of GVHD. In some embodiments, the ACK inhibitor compound(e.g., an ITK or BTK inhibitor, such as for example ibrutinib) is usedas a prophylactic and is administered continuously to subjects with apropensity to develop GVHD (e.g., allogeneic transplant recipients). Insome embodiments, the ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as for example ibrutinib) is administered to anindividual during or as soon as possible after the development of GVHD.In some embodiments, the administration of the ACK inhibitor compound(e.g., an ITK or BTK inhibitor, such as for example ibrutinib) isinitiated within the first 48 hours of the onset of the symptoms, withinthe first 6 hours of the onset of the symptoms, or within 3 hours of theonset of the symptoms. In some embodiments, the initial administrationof the ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such asfor example ibrutinib) is via any route practical, such as, for example,an intravenous injection, a bolus injection, infusion over 5 minutes toabout 5 hours, a pill, a capsule, a tablet, a transdermal patch, buccaldelivery, and the like, or combination thereof. The ACK inhibitorcompound (e.g., an ITK or BTK inhibitor, such as for example ibrutinib)should be administered as soon as is practicable after the onset of adisorder is detected or suspected, and for a length of time necessaryfor the treatment of the disease, such as, for example, from about 1month to about 3 months. The length of treatment can vary for eachsubject, and the length can be determined using the known criteria. Insome embodiments, the ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as for example ibrutinib) is administered for at least 2weeks, between about 1 month to about 5 years, or from about 1 month toabout 3 years.

Therapeutically effective amounts will depend on the severity and courseof the disorder, previous therapy, the patient's health status, weight,and response to the drugs, and the judgment of the treating physician.Prophylactically effective amounts depend on the patient's state ofhealth, weight, the severity and course of the disease, previoustherapy, response to the drugs, and the judgment of the treatingphysician.

In some embodiments, the ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as for example ibrutinib) is administered to the patienton a regular basis, e.g., three times a day, two times a day, once aday, every other day or every 3 days. In other embodiments, the ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as for exampleibrutinib) is administered to the patient on an intermittent basis,e.g., twice a day followed by once a day followed by three times a day;or the first two days of every week; or the first, second and third dayof a week. In some embodiments, intermittent dosing is as effective asregular dosing. In further or alternative embodiments, the ACK inhibitorcompound (e.g., an ITK or BTK inhibitor, such as for example ibrutinib)is administered only when the patient exhibits a particular symptom,e.g., the onset of pain, or the onset of a fever, or the onset of aninflammation, or the onset of a skin disorder. Dosing schedules of eachcompound may depend on the other or may be independent of the other.

In the case wherein the patient's condition does not improve, upon thedoctor's discretion the compounds may be administered chronically, thatis, for an extended period of time, including throughout the duration ofthe patient's life in order to ameliorate or otherwise control or limitthe symptoms of the patient's disorder.

In the case wherein the patient's status does improve, upon the doctor'sdiscretion the compounds may be given continuously; alternatively, thedose of drug being administered may be temporarily reduced ortemporarily suspended for a certain length of time (i.e., a “drugholiday”). The length of the drug holiday can vary between 2 days and 1year, including by way of example only, 2 days, 3 days, 4 days, 5 days,6 days, 7 days, 10 days, 12 days, 15 days, 20 days, 28 days, 35 days, 50days, 70 days, 100 days, 120 days, 150 days, 180 days, 200 days, 250days, 280 days, 300 days, 320 days, 350 days, or 365 days. The dosereduction during a drug holiday may be from 10%-100%, including, by wayof example only, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%,65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100%.

Once improvement of the patient's conditions has occurred, a maintenanceregimen is administered if necessary. Subsequently, the dosage or thefrequency of administration, or both, of the ACK inhibitor compound(e.g., an ITK or BTK inhibitor, such as for example ibrutinib) can bereduced, as a function of the symptoms, to a level at which theindividual's improved condition is retained. Individuals can, however,require intermittent treatment on a long-term basis upon any recurrenceof symptoms.

The amount of the ACK inhibitor compound (e.g., an ITK or BTK inhibitor,such as for example ibrutinib) will vary depending upon factors such asthe particular compound, disorder and its severity, the identity (e.g.,weight) of the subject or host in need of treatment, and is determinedaccording to the particular circumstances surrounding the case,including, e.g., the specific agents being administered, the routes ofadministration, and the subject or host being treated. In general,however, doses employed for adult human treatment will typically be inthe range of 0.02-5000 mg per day, or from about 1-1500 mg per day. Thedesired dose may be presented in a single dose or as divided dosesadministered simultaneously (or over a short period of time) or atappropriate intervals, for example as two, three, four or more sub-dosesper day.

In some embodiments, the therapeutic amount of the ACK inhibitor (e.g.,an ITK or BTK inhibitor, such as for example ibrutinib) is from 100mg/day up to, and including, 2000 mg/day. In some embodiments, theamount of the ACK inhibitor (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib) is from 140 mg/day up to, and including, 840 mg/day.In some embodiments, the amount of the ACK inhibitor (e.g., an ITK orBTK inhibitor, such as for example ibrutinib) is from 420 mg/day up to,and including, 840 mg/day. In some embodiments, the amount of the ACKinhibitor (e.g., an ITK or BTK inhibitor, such as for example ibrutinib)is about 40 mg/day. In some embodiments, the amount of the ACK inhibitor(e.g., an ITK or BTK inhibitor, such as for example ibrutinib) is about140 mg/day. In some embodiments, the amount of the ACK inhibitor (e.g.,an ITK or BTK inhibitor, such as for example ibrutinib) is about 280mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 420mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 560mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 700mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 840mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 980mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 1120mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 1260mg/day. In some embodiments, the amount of the ACK inhibitor (e.g., anITK or BTK inhibitor, such as for example ibrutinib) is about 1400mg/day. In some embodiments, a compound of Formula (A) is administeredat a dosage of between about 0.1 mg/kg per day to about 100 mg/kg perday.

In some embodiments, the dosage of the ACK inhibitor (e.g., an ITK orBTK inhibitor, such as for example ibrutinib) is escalated over time. Insome embodiments, the dosage of the ACK inhibitor (e.g., an ITK or BTKinhibitor, such as for example ibrutinib) is escalated, for example,from at or about 1.25 mg/kg/day to at or about 12.5 mg/kg/day over apredetermined period of time. In some embodiments the predeterminedperiod of time is over 1 month, over 2 months, over 3 months, over 4months, over 5 months, over 6 months, over 7 months, over 8 months, over9 months, over 10 months, over 11 months, over 12 months, over 18months, over 24 months or longer.

The ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib) may be formulated into unit dosage forms suitable forsingle administration of precise dosages. In unit dosage form, theformulation is divided into unit doses containing appropriate quantitiesof one or both compounds. The unit dosage may be in the form of apackage containing discrete quantities of the formulation. Non-limitingexamples are packaged tablets or capsules, and powders in vials orampoules. Aqueous suspension compositions can be packaged in single-dosenon-reclosable containers. Alternatively, multiple-dose reclosablecontainers can be used, in which case it is typical to include apreservative in the composition. By way of example only, formulationsfor parenteral injection may be presented in unit dosage form, whichinclude, but are not limited to ampoules, or in multi-dose containers,with an added preservative.

It is understood that a medical professional will determine the dosageregimen in accordance with a variety of factors. These factors includethe severity of GVHD in the subject, as well as the age, weight, sex,diet, and medical condition of the subject.

Compounds

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administering tothe patient a composition comprising a therapeutically-effective amountof an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib).

Further described herein are methods of treating a patient foralleviation of a bone marrow mediated disease, with alleviation ofconsequently developed graft versus host disease (GVHD), comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, wherein a therapeutically effective amount of an ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as for exampleibrutinib) is administered prior to or concurrently with the allogeneichematopoietic stem cells and/or allogeneic T-cells.

In the following description of irreversible BTK compounds suitable foruse in the methods described herein, definitions of referred-to standardchemistry terms may be found in reference works (if not otherwisedefined herein), including Carey and Sundberg “Advanced OrganicChemistry 4th Ed.” Vols. A (2000) and B (2001), Plenum Press, New York.Unless otherwise indicated, conventional methods of mass spectroscopy,NMR, HPLC, protein chemistry, biochemistry, recombinant DNA techniquesand pharmacology, within the ordinary skill of the art are employed. Inaddition, nucleic acid and amino acid sequences for BTK (e.g., humanBTK) are known in the art as disclosed in, e.g., U.S. Pat. No.6,326,469. Unless specific definitions are provided, the nomenclatureemployed in connection with, and the laboratory procedures andtechniques of, analytical chemistry, synthetic organic chemistry, andmedicinal and pharmaceutical chemistry described herein are those knownin the art. Standard techniques can be used for chemical syntheses,chemical analyses, pharmaceutical preparation, formulation, anddelivery, and treatment of patients.

The BTK inhibitor compounds described herein are selective for BTK andkinases having a cysteine residue in an amino acid sequence position ofthe tyrosine kinase that is homologous to the amino acid sequenceposition of cysteine 481 in BTK. Generally, an irreversible inhibitorcompound of BTK used in the methods described herein is identified orcharacterized in an in vitro assay, e.g., an acellular biochemical assayor a cellular functional assay. Such assays are useful to determine anin vitro IC₅₀ for an irreversible BTK inhibitor compound.

For example, an acellular kinase assay can be used to determine BTKactivity after incubation of the kinase in the absence or presence of arange of concentrations of a candidate irreversible BTK inhibitorcompound. If the candidate compound is in fact an irreversible BTKinhibitor, BTK kinase activity will not be recovered by repeat washingwith inhibitor-free medium. See, e.g., J. B. Smaill, et al. (1999), J.Med. Chem. 42(10):1803-1815. Further, covalent complex formation betweenBTK and a candidate irreversible BTK inhibitor is a useful indicator ofirreversible inhibition of BTK that can be readily determined by anumber of methods known in the art (e.g., mass spectrometry). Forexample, some irreversible BTK-inhibitor compounds can form a covalentbond with Cys 481 of BTK (e.g., via a Michael reaction).

Cellular functional assays for BTK inhibition include measuring one ormore cellular endpoints in response to stimulating a BTK-mediatedpathway in a cell line (e.g., BCR activation in Ramos cells) in theabsence or presence of a range of concentrations of a candidateirreversible BTK inhibitor compound. Useful endpoints for determining aresponse to BCR activation include, e.g., autophosphorylation of BTK,phosphorylation of a BTK target protein (e.g., PLC-γ), and cytoplasmiccalcium flux.

High-throughput assays for many acellular biochemical assays (e.g.,kinase assays) and cellular functional assays (e.g., calcium flux) arewell known to those of ordinary skill in the art. In addition, highthroughput screening systems are commercially available (see, e.g.,Zymark Corp., Hopkinton, Mass.; Air Technical Industries, Mentor, OH;Beckman Instruments, Inc. Fullerton, Calif.; Precision Systems, Inc.,Natick, Mass., etc.). These systems typically automate entire proceduresincluding all sample and reagent pipetting, liquid dispensing, timedincubations, and final readings of the microplate in detector(s)appropriate for the assay. Automated systems thereby allow theidentification and characterization of a large number of irreversibleBTK compounds without undue effort.

In some embodiments, the BTK inhibitor is selected from the groupconsisting of a small organic molecule, a macromolecule, a peptide or anon-peptide.

In some embodiments, the BTK inhibitor provided herein is a reversibleor irreversible inhibitor. In certain embodiments, the BTK inhibitor isan irreversible inhibitor.

In some embodiments, the irreversible BTK inhibitor forms a covalentbond with a cysteine sidechain of a Bruton's tyrosine kinase, a Bruton'styrosine kinase homolog, or a BTK tyrosine kinase cysteine homolog.

Irreversible BTK inhibitor compounds can be used for the manufacture ofa medicament for treating any of the foregoing conditions (e.g.,autoimmune diseases, inflammatory diseases, allergy disorders, B-cellproliferative disorders, or thromboembolic disorders).

In some embodiments, the irreversible BTK inhibitor compound used forthe methods described herein inhibits BTK or a BTK homolog kinaseactivity with an in vitro IC₅₀ of less than 10 μM (e.g., less than 1 μM,less than 0.5 μM, less than 0.4 μM, less than 0.3 μM, less than 0.1,less than 0.08 μM, less than 0.06 μM, less than 0.05 μM, less than 0.04μM, less than 0.03 μM, less than less than 0.02 μM, less than 0.01, lessthan 0.008 μM, less than 0.006 μM, less than 0.005 μM, less than 0.004μM, less than 0.003 μM, less than less than 0.002 μM, less than 0.001,less than 0.00099 μM, less than 0.00098 μM, less than 0.00097 μM, lessthan 0.00096 μM, less than 0.00095 μM, less than 0.00094 μM, less than0.00093 μM, less than 0.00092, or less than 0.00090 μM).

In some embodiments, the irreversible BTK inhibitor compound is selectedfrom among ibrutinib (PCI-32765), PCI-45292, PCI-45466, AVL-101,AVL-291, AVL-292, or ONO-WG-37. In some embodiments, the irreversibleBTK inhibitor compound is ibrutinib.

In one embodiment, the irreversible BTK inhibitor compound selectivelyand irreversibly inhibits an activated form of its target tyrosinekinase (e.g., a phosphorylated form of the tyrosine kinase). Forexample, activated BTK is transphosphorylated at tyrosine 551. Thus, inthese embodiments the irreversible BTK inhibitor inhibits the targetkinase in cells only once the target kinase is activated by thesignaling events.

In other embodiments, the BTK inhibitor used in the methods describeherein has the structure of any of Formula (A). Also described hereinare pharmaceutically acceptable salts, pharmaceutically acceptablesolvates, pharmaceutically active metabolites, and pharmaceuticallyacceptable prodrugs of such compounds. Pharmaceutical compositions thatinclude at least one such compound or a pharmaceutically acceptablesalt, pharmaceutically acceptable solvate, pharmaceutically activemetabolite or pharmaceutically acceptable prodrug of such compound, areprovided.

Definition of standard chemistry terms are found in reference works,including Carey and Sundberg “ADVANCED ORGANIC CHEMISTRY 4 ED.” Vols. A(2000) and B (2001), Plenum Press, New York. Unless otherwise indicated,conventional methods of mass spectroscopy, NMR, HPLC, protein chemistry,biochemistry, recombinant DNA techniques and pharmacology, within theskill of the art are employed. Unless specific definitions are provided,the nomenclature employed in connection with, and the laboratoryprocedures and techniques of, analytical chemistry, synthetic organicchemistry, and medicinal and pharmaceutical chemistry described hereinare those known in the art. Standard techniques are optionally used forchemical syntheses, chemical analyses, pharmaceutical preparation,formulation, and delivery, and treatment of patients. Standardtechniques are optionally used for recombinant DNA, oligonucleotidesynthesis, and tissue culture and transformation (e.g., electroporation,lipofection). Reactions and purification techniques are performed usingdocumented methodologies or as described herein.

It is to be understood that the methods and compositions describedherein are not limited to the particular methodology, protocols, celllines, constructs, and reagents described herein and as such optionallyvary. It is also to be understood that the terminology used herein isfor the purpose of describing particular embodiments only, and is notintended to limit the scope of the methods and compositions describedherein, which will be limited only by the appended claims.

Unless stated otherwise, the terms used for complex moieties (i.e.,multiple chains of moieties) are to be read equivalently either fromleft to right or right to left. For example, the groupalkylenecycloalkylene refers both to an alkylene group followed by acycloalkylene group or as a cycloalkylene group followed by an alkylenegroup.

The suffix “ene” appended to a group indicates that such a group is adiradical. By way of example only, a methylene is a diradical of amethyl group, that is, it is a —CH₂— group; and an ethylene is adiradical of an ethyl group, i.e., —CH₂CH₂—.

An “alkyl” group refers to an aliphatic hydrocarbon group. The alkylmoiety includes a “saturated alkyl” group, which means that it does notcontain any alkene or alkyne moieties. The alkyl moiety also includes an“unsaturated alkyl” moiety, which means that it contains at least onealkene or alkyne moiety. An “alkene” moiety refers to a group that hasat least one carbon-carbon double bond, and an “alkyne” moiety refers toa group that has at least one carbon-carbon triple bond. The alkylmoiety, whether saturated or unsaturated, includes branched, straightchain, or cyclic moieties. Depending on the structure, an alkyl groupincludes a monoradical or a diradical (i.e., an alkylene group), and ifa “lower alkyl” having 1 to 6 carbon atoms.

As used herein, C₁-C_(x) includes C₁-C₂, C₁-C₃ . . . C₁-C_(x).

The “alkyl” moiety optionally has 1 to 10 carbon atoms (whenever itappears herein, a numerical range such as “1 to 10” refers to eachinteger in the given range; e.g., “1 to 10 carbon atoms” means that thealkyl group is selected from a moiety having 1 carbon atom, 2 carbonatoms, 3 carbon atoms, etc., up to and including 10 carbon atoms,although the present definition also covers the occurrence of the term“alkyl” where no numerical range is designated). The alkyl group of thecompounds described herein may be designated as “C₁-C₄ alkyl” or similardesignations. By way of example only, “C₁-C₄ alkyl” indicates that thereare one to four carbon atoms in the alkyl chain, i.e., the alkyl chainis selected from among methyl, ethyl, propyl, iso-propyl, n-butyl,iso-butyl, sec-butyl, and t-butyl. Thus C₁-C₄ alkyl includes C₁-C₂ alkyland C₁-C₃ alkyl. Alkyl groups are optionally substituted orunsubstituted. Typical alkyl groups include, but are in no way limitedto, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl,pentyl, hexyl, ethenyl, propenyl, butenyl, cyclopropyl, cyclobutyl,cyclopentyl, cyclohexyl, and the like.

The term “alkenyl” refers to a type of alkyl group in which the firsttwo atoms of the alkyl group form a double bond that is not part of anaromatic group. That is, an alkenyl group begins with the atoms—C(R)═C(R)—R, wherein R refers to the remaining portions of the alkenylgroup, which are either the same or different. The alkenyl moiety isoptionally branched, straight chain, or cyclic (in which case, it isalso known as a “cycloalkenyl” group). Depending on the structure, analkenyl group includes a monoradical or a diradical (i.e., an alkenylenegroup). Alkenyl groups are optionally substituted. Non-limiting examplesof an alkenyl group include —CH═CH₂, —C(CH₃)═CH₂, —CH═CHCH₃,—C(CH₃)═CHCH₃. Alkenylene groups include, but are not limited to,—CH═CH—, —C(CH₃)═CH—, —CH═CHCH₂—, —CH═CHCH₂CH₂— and —C(CH₃)═CHCH₂—.Alkenyl groups optionally have 2 to 10 carbons, and if a “lower alkenyl”having 2 to 6 carbon atoms.

The term “alkynyl” refers to a type of alkyl group in which the firsttwo atoms of the alkyl group form a triple bond. That is, an alkynylgroup begins with the atoms —C≡C—R, wherein R refers to the remainingportions of the alkynyl group, which is either the same or different.The “R” portion of the alkynyl moiety may be branched, straight chain,or cyclic. Depending on the structure, an alkynyl group includes amonoradical or a diradical (i.e., an alkynylene group). Alkynyl groupsare optionally substituted. Non-limiting examples of an alkynyl groupinclude, but are not limited to, —C≡CH, —C≡CCH₃, —C≡CCH₂CH₃, and—C≡CCH₂. Alkynyl groups optionally have 2 to 10 carbons, and if a “loweralkynyl” having 2 to 6 carbon atoms.

An “alkoxy” group refers to a (alkyl)O— group, where alkyl is as definedherein.

“Hydroxyalkyl” refers to an alkyl radical, as defined herein,substituted with at least one hydroxy group. Non-limiting examples of ahydroxyalkyl include, but are not limited to, hydroxymethyl,2-hydroxyethyl, 2-hydroxypropyl, 3-hydroxypropyl,1-(hydroxymethyl)-2-methylpropyl, 2-hydroxybutyl, 3-hydroxybutyl,4-hydroxybutyl, 2,3-dihydroxypropyl, 1-(hydroxymethyl)-2-hydroxyethyl,2,3-dihydroxybutyl, 3,4-dihydroxybutyl and2-(hydroxymethyl)-3-hydroxypropyl.

“Alkoxyalkyl” refers to an alkyl radical, as defined herein, substitutedwith an alkoxy group, as defined herein.

The term “alkylamine” refers to the —N(alkyl)_(x)H_(y) group, where xand y are selected from among x=1, y=1 and x=2, y=0. When x=2, the alkylgroups, taken together with the N atom to which they are attached,optionally form a cyclic ring system.

“Alkylaminoalkyl” refers to an alkyl radical, as defined herein,substituted with an alkylamine, as defined herein.

“Hydroxyalkylaminoalkyl” refers to an alkyl radical, as defined herein,substituted with an alkylamine, and alkylhydroxy, as defined herein.

“Alkoxyalkylaminoalkyl” refers to an alkyl radical, as defined herein,substituted with an alkylamine and substituted with an alkylalkoxy, asdefined herein.

An “amide” is a chemical moiety with the formula —C(O)NHR or —NHC(O)R,where R is selected from among alkyl, cycloalkyl, aryl, heteroaryl(bonded through a ring carbon) and heteroalicyclic (bonded through aring carbon). In some embodiments, an amide moiety forms a linkagebetween an amino acid or a peptide molecule and a compound describedherein, thereby forming a prodrug. Any amine, or carboxyl side chain onthe compounds described herein can be amidified. The procedures andspecific groups to make such amides are found in sources such as Greeneand Wuts, Protective Groups in Organic Synthesis, 3^(rd) Ed., John Wiley& Sons, New York, N.Y., 1999, which is incorporated herein by referencefor this disclosure.

The term “ester” refers to a chemical moiety with formula —COOR, where Ris selected from among alkyl, cycloalkyl, aryl, heteroaryl (bondedthrough a ring carbon) and heteroalicyclic (bonded through a ringcarbon). Any hydroxy, or carboxyl side chain on the compounds describedherein can be esterified. The procedures and specific groups to makesuch esters are found in sources such as Greene and Wuts, ProtectiveGroups in Organic Synthesis, 3^(rd) Ed., John Wiley & Sons, New York,N.Y., 1999, which is incorporated herein by reference for thisdisclosure.

As used herein, the term “ring” refers to any covalently closedstructure. Rings include, for example, carbocycles (e.g., aryls andcycloalkyls), heterocycles (e.g., heteroaryls and non-aromaticheterocycles), aromatics (e.g. aryls and heteroaryls), and non-aromatics(e.g., cycloalkyls and non-aromatic heterocycles). Rings can beoptionally substituted. Rings can be monocyclic or polycyclic.

As used herein, the term “ring system” refers to one, or more than onering.

The term “membered ring” can embrace any cyclic structure. The term“membered” is meant to denote the number of skeletal atoms thatconstitute the ring. Thus, for example, cyclohexyl, pyridine, pyran andthiopyran are 6-membered rings and cyclopentyl, pyrrole, furan, andthiophene are 5-membered rings.

The term “fused” refers to structures in which two or more rings shareone or more bonds.

The term “carbocyclic” or “carbocycle” refers to a ring wherein each ofthe atoms forming the ring is a carbon atom. Carbocycle includes aryland cycloalkyl. The term thus distinguishes carbocycle from heterocycle(“heterocyclic”) in which the ring backbone contains at least one atomwhich is different from carbon (i.e. a heteroatom). Heterocycle includesheteroaryl and heterocycloalkyl. Carbocycles and heterocycles can beoptionally substituted.

The term “aromatic” refers to a planar ring having a delocalized7c-electron system containing 4n+2 π electrons, where n is an integer.Aromatic rings can be formed from five, six, seven, eight, nine, or morethan nine atoms. Aromatics can be optionally substituted. The term“aromatic” includes both carbocyclic aryl (e.g., phenyl) andheterocyclic aryl (or “heteroaryl” or “heteroaromatic”) groups (e.g.,pyridine). The term includes monocyclic or fused-ring polycyclic (i.e.,rings which share adjacent pairs of carbon atoms) groups.

As used herein, the term “aryl” refers to an aromatic ring wherein eachof the atoms forming the ring is a carbon atom. Aryl rings can be formedby five, six, seven, eight, nine, or more than nine carbon atoms. Arylgroups can be optionally substituted. Examples of aryl groups include,but are not limited to phenyl, naphthalenyl, phenanthrenyl, anthracenyl,fluorenyl, and indenyl. Depending on the structure, an aryl group can bea monoradical or a diradical (i.e., an arylene group).

An “aryloxy” group refers to an (aryl)O— group, where aryl is as definedherein.

The term “carbonyl” as used herein refers to a group containing a moietyselected from the group consisting of —C(O)—, —S(O)—, —S(O)₂—, and—C(S)—, including, but not limited to, groups containing a least oneketone group, and/or at least one aldehyde group, and/or at least oneester group, and/or at least one carboxylic acid group, and/or at leastone thioester group. Such carbonyl groups include ketones, aldehydes,carboxylic acids, esters, and thioesters. In some embodiments, suchgroups are a part of linear, branched, or cyclic molecules.

The term “cycloalkyl” refers to a monocyclic or polycyclic radical thatcontains only carbon and hydrogen, and is optionally saturated,partially unsaturated, or fully unsaturated. Cycloalkyl groups includegroups having from 3 to 10 ring atoms. Illustrative examples ofcycloalkyl groups include the following moieties:

and the like. Depending on the structure, a cycloalkyl group is either amonoradical or a diradical (e.g., an cycloalkylene group), and if a“lower cycloalkyl” having 3 to 8 carbon atoms.

“Cycloalkylalkyl” means an alkyl radical, as defined herein, substitutedwith a cycloalkyl group. Non-limiting cycloalkylalkyl groups includecyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl,cyclohexylmethyl, and the like.

The term “heterocycle” refers to heteroaromatic and heteroalicyclicgroups containing one to four heteroatoms each selected from O, S and N,wherein each heterocyclic group has from 4 to 10 atoms in its ringsystem, and with the proviso that the ring of said group does notcontain two adjacent O or S atoms. Herein, whenever the number of carbonatoms in a heterocycle is indicated (e.g., C₁-C₆ heterocycle), at leastone other atom (the heteroatom) must be present in the ring.Designations such as “C₁-C₆ heterocycle” refer only to the number ofcarbon atoms in the ring and do not refer to the total number of atomsin the ring. It is understood that the heterocylic ring can haveadditional heteroatoms in the ring. Designations such as “4-6 memberedheterocycle” refer to the total number of atoms that are contained inthe ring (i.e., a four, five, or six membered ring, in which at leastone atom is a carbon atom, at least one atom is a heteroatom and theremaining two to four atoms are either carbon atoms or heteroatoms). Inheterocycles that have two or more heteroatoms, those two or moreheteroatoms can be the same or different from one another. Heterocyclescan be optionally substituted. Binding to a heterocycle can be at aheteroatom or via a carbon atom. Non-aromatic heterocyclic groupsinclude groups having only 4 atoms in their ring system, but aromaticheterocyclic groups must have at least 5 atoms in their ring system. Theheterocyclic groups include benzo-fused ring systems. An example of a4-membered heterocyclic group is azetidinyl (derived from azetidine). Anexample of a 5-membered heterocyclic group is thiazolyl. An example of a6-membered heterocyclic group is pyridyl, and an example of a10-membered heterocyclic group is quinolinyl. Examples of non-aromaticheterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl,tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl,tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino,thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl,homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl,thiazepinyl, 1,2,3,6-tetrahydropyridinyl, 2-pyrrolinyl, 3-pyrrolinyl,indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl,pyrazolinyl, dithianyl, dithiolanyl, dihydropyranyl, dihydrothienyl,dihydrofuranyl, pyrazolidinyl, imidazolinyl, imidazolidinyl,3-azabicyclo[3.1.0]hexanyl, 3-azabicyclo[4.1.0]heptanyl, 3H-indolyl andquinolizinyl. Examples of aromatic heterocyclic groups are pyridinyl,imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl,furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl,quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl,cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl,triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl,furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl,benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, andfuropyridinyl. The foregoing groups, as derived from the groups listedabove, are optionally C-attached or N-attached where such is possible.For instance, a group derived from pyrrole includes pyrrol-1-yl(N-attached) or pyrrol-3-yl (C-attached). Further, a group derived fromimidazole includes imidazol-1-yl or imidazol-3-yl (both N-attached) orimidazol-2-yl, imidazol-4-yl or imidazol-5-yl (all C-attached). Theheterocyclic groups include benzo-fused ring systems and ring systemssubstituted with one or two oxo (═O) moieties such as pyrrolidin-2-one.Depending on the structure, a heterocycle group can be a monoradical ora diradical (i.e., a heterocyclene group).

The terms “heteroaryl” or, alternatively, “heteroaromatic” refers to anaromatic group that includes one or more ring heteroatoms selected fromnitrogen, oxygen and sulfur. An N-containing “heteroaromatic” or“heteroaryl” moiety refers to an aromatic group in which at least one ofthe skeletal atoms of the ring is a nitrogen atom. Illustrative examplesof heteroaryl groups include the following moieties:

and the like. Depending on the structure, a heteroaryl group can be amonoradical or a diradical (i.e., a heteroarylene group).

As used herein, the term “non-aromatic heterocycle”, “heterocycloalkyl”or “heteroalicyclic” refers to a non-aromatic ring wherein one or moreatoms forming the ring is a heteroatom. A “non-aromatic heterocycle” or“heterocycloalkyl” group refers to a cycloalkyl group that includes atleast one heteroatom selected from nitrogen, oxygen and sulfur. In someembodiments, the radicals are fused with an aryl or heteroaryl.Heterocycloalkyl rings can be formed by three, four, five, six, seven,eight, nine, or more than nine atoms. Heterocycloalkyl rings can beoptionally substituted. In certain embodiments, non-aromaticheterocycles contain one or more carbonyl or thiocarbonyl groups suchas, for example, oxo- and thio-containing groups. Examples ofheterocycloalkyls include, but are not limited to, lactams, lactones,cyclic imides, cyclic thioimides, cyclic carbamates,tetrahydrothiopyran, 4H-pyran, tetrahydropyran, piperidine, 1,3-dioxin,1,3-dioxane, 1,4-dioxin, 1,4-dioxane, piperazine, 1,3-oxathiane,1,4-oxathiin, 1,4-oxathiane, tetrahydro-1,4-thiazine, 2H-1,2-oxazine,maleimide, succinimide, barbituric acid, thiobarbituric acid,dioxopiperazine, hydantoin, dihydrouracil, morpholine, trioxane,hexahydro-1,3,5-triazine, tetrahydrothiophene, tetrahydrofuran,pyrroline, pyrrolidine, pyrrolidone, pyrrolidione, pyrazoline,pyrazolidine, imidazoline, imidazolidine, 1,3-dioxole, 1,3-dioxolane,1,3-dithiole, 1,3-dithiolane, isoxazoline, isoxazolidine, oxazoline,oxazolidine, oxazolidinone, thiazoline, thiazolidine, and1,3-oxathiolane. Illustrative examples of heterocycloalkyl groups, alsoreferred to as non-aromatic heterocycles, include:

and the like. The term heteroalicyclic also includes all ring forms ofthe carbohydrates, including but not limited to the monosaccharides, thedisaccharides and the oligosaccharides. Depending on the structure, aheterocycloalkyl group can be a monoradical or a diradical (i.e., aheterocycloalkylene group).

The term “halo” or, alternatively, “halogen” or “halide” means fluoro,chloro, bromo, and iodo.

The term “haloalkyl,” refers to alkyl structures in which at least onehydrogen is replaced with a halogen atom. In certain embodiments inwhich two or more hydrogen atoms are replaced with halogen atoms, thehalogen atoms are all the same as one another. In other embodiments inwhich two or more hydrogen atoms are replaced with halogen atoms, thehalogen atoms are not all the same as one another.

The term “fluoroalkyl,” as used herein, refers to alkyl group in whichat least one hydrogen is replaced with a fluorine atom. Examples offluoroalkyl groups include, but are not limited to, —CF₃, —CH₂CF₃,—CF₂CF₃, —CH₂CH₂CF₃ and the like.

As used herein, the term “heteroalkyl” refers to optionally substitutedalkyl radicals in which one or more skeletal chain atoms is aheteroatom, e.g., oxygen, nitrogen, sulfur, silicon, phosphorus orcombinations thereof. The heteroatom(s) are placed at any interiorposition of the heteroalkyl group or at the position at which theheteroalkyl group is attached to the remainder of the molecule. Examplesinclude, but are not limited to, —CH₂—O—CH₃, —CH₂—CH₂—O—CH₃,—CH₂—NH—CH₃, —CH₂—CH₂—NH—CH₃, —CH₂—N(CH₃)—CH₃, —CH₂—CH₂—NH—CH₃,—CH₂—CH₂—N(CH₃)—CH₃, —CH₂—S—CH₂— CH₃, —CH₂—CH₂, —S(O)—CH₃,—CH₂—CH₂—S(O)₂—CH₃, —CH═CH—O—CH₃, —Si(CH₃)₃, —CH₂—CH═N—OCH₃, and—CH═CH—N(CH₃)—CH₃. In addition, in some embodiments, up to twoheteroatoms are consecutive, such as, by way of example, —CH₂—NH—OCH₃and —CH₂—O—Si(CH₃)₃.

The term “heteroatom” refers to an atom other than carbon or hydrogen.Heteroatoms are typically independently selected from among oxygen,sulfur, nitrogen, silicon and phosphorus, but are not limited to theseatoms. In embodiments in which two or more heteroatoms are present, thetwo or more heteroatoms can all be the same as one another, or some orall of the two or more heteroatoms can each be different from theothers.

The term “bond” or “single bond” refers to a chemical bond between twoatoms, or two moieties when the atoms joined by the bond are consideredto be part of larger substructure.

The term “moiety” refers to a specific segment or functional group of amolecule. Chemical moieties are often recognized chemical entitiesembedded in or appended to a molecule.

A “thioalkoxy” or “alkylthio” group refers to a —S-alkyl group.

A “SH” group is also referred to either as a thiol group or a sulfhydrylgroup.

The term “optionally substituted” or “substituted” means that thereferenced group may be substituted with one or more additional group(s)individually and independently selected from alkyl, cycloalkyl, aryl,heteroaryl, heteroalicyclic, hydroxy, alkoxy, aryloxy, alkylthio,arylthio, alkylsulfoxide, arylsulfoxide, alkylsulfone, arylsulfone,cyano, halo, acyl, nitro, haloalkyl, fluoroalkyl, amino, including mono-and di-substituted amino groups, and the protected derivatives thereof.By way of example an optional substituents may be L_(s)R_(s), whereineach L_(s) is independently selected from a bond, —O—, —C(═O)—, —S—,—S(═O)—, —S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, S(═O)₂NH—, —NHS(═O)₂,—OC(O)NH—, —NHC(O)O—, -(substituted or unsubstituted C₁-C₆ alkyl), or-(substituted or unsubstituted C₂-C₆ alkenyl); and each R_(s) isindependently selected from H, (substituted or unsubstitutedC₁-C₄alkyl), (substituted or unsubstituted C₃-C₆cycloalkyl), heteroaryl,or heteroalkyl. The protecting groups that form the protectivederivatives of the above substituents include those found in sourcessuch as Greene and Wuts, above.

ACK Inhibitor Compounds

Described herein are methods of preventing the occurrence of graftversus host disease (GVHD) or reducing the severity of GVHD occurrencein a patient requiring cell transplantation comprising administering tothe patient a composition comprising a therapeutically-effective amountof an ACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as,for example, ibrutinib).

Further described herein are methods of treating a patient foralleviation of a bone marrow mediated disease, with alleviation ofconsequently developed graft versus host disease (GVHD), comprisingadministering to the patient allogeneic hematopoietic stem cells and/orallogeneic T-cells, wherein a therapeutically effective amount of an ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as, for example,ibrutinib) is administered prior to or concurrently with the allogeneichematopoietic stem cells and/or allogeneic T-cells.

The ACK inhibitor compounds described herein are selective for kinaseshaving an accessible cysteine that is able to form a covalent bond witha Michael acceptor moiety on the inhibitor compound. In someembodiments, the cysteine residue is accessible or becomes accessiblewhen the binding site moiety of the irreversible inhibitor binds to thekinase. That is, the binding site moiety of the irreversible inhibitorbinds to an active site of the ACK and the Michael acceptor moiety ofirreversible inhibitor gains access (in one embodiment the step ofbinding leads to a conformational change in the ACK, thus exposing thecysteine) or is otherwise exposed to the cysteine residue of the ACK; asa result a covalent bond is formed between the “S” of the cysteineresidue and the Michael acceptor of the irreversible inhibitor.Consequently, the binding site moiety of the irreversible inhibitorremains bound or otherwise blocks the active site of the ACK.

In some embodiments, the ACK is BTK, a homolog of BTK or a tyrosinekinase having a cysteine residue in an amino acid sequence position thatis homologous to the amino acid sequence position of cysteine 481 inBTK. In some embodiments, the ACK is ITK. In some embodiments, the ACKis HER4. Inhibitor compounds described herein include a Michael acceptormoiety, a binding site moiety and a linker that links the binding sitemoiety and the Michael acceptor moiety (and in some embodiments, thestructure of the linker provides a conformation, or otherwise directsthe Michael acceptor moiety, so as to improve the selectivity of theirreversible inhibitor for a particular ACK). In some embodiments, theACK inhibitor inhibits ITK and BTK.

In some embodiments, the ACK inhibitor is a compound of Formula (A):

wherein

-   -   A is independently selected from N or CR₅;    -   R₁ is H, L₂-(substituted or unsubstituted alkyl),        L₂-(substituted or unsubstituted cycloalkyl), L₂-(substituted or        unsubstituted alkenyl), L₂-(substituted or unsubstituted        cycloalkenyl), L₂-(substituted or unsubstituted heterocycle),        L₂-(substituted or unsubstituted heteroaryl), or L₂-(substituted        or unsubstituted aryl), where L₂ is a bond, O, S, —S(═O),        —S(═O)₂, C(═O), -(substituted or unsubstituted C₁-C₆ alkyl), or        -(substituted or unsubstituted C₂-C₆ alkenyl);    -   R₂ and R₃ are independently selected from H, lower alkyl and        substituted lower alkyl;    -   R₄ is L₃-X-L₄-G, wherein,        -   L₃ is optional, and when present is a bond, optionally            substituted or unsubstituted alkyl, optionally substituted            or unsubstituted cycloalkyl, optionally substituted or            unsubstituted alkenyl, optionally substituted or            unsubstituted alkynyl; X is optional, and when present is a            bond, O, —C(═O), S, —S(═O), —S(═O)₂, —NH, —NR₉, —NHC(O),            —C(O)NH, —NR₉C(O), —C(O)NR₉, —S(═O)₂NH, —NHS(═O)₂,            —S(═O)₂NR₉—, —NR₉S(═O)₂, —OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—,            —NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—, heteroaryl,            aryl, —NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—,            —OC(═NR₁₁)—, or —C(═NR₁₁)O—;        -   L₄ is optional, and when present is a bond, substituted or            unsubstituted alkyl, substituted or unsubstituted            cycloalkyl, substituted or unsubstituted alkenyl,            substituted or unsubstituted alkynyl, substituted or            unsubstituted aryl, substituted or unsubstituted heteroaryl,            substituted or unsubstituted heterocycle;        -   or L₃, X and L₄ taken together form a nitrogen containing            heterocyclic ring;

G is

wherein,

-   -   -   R₆, R₇ and R₈ are independently selected from among H, lower            alkyl or substituted lower alkyl, lower heteroalkyl or            substituted lower heteroalkyl, substituted or unsubstituted            lower cycloalkyl, and substituted or unsubstituted lower            heterocycloalkyl;

    -   R₅ is H, halogen, -L₆-(substituted or unsubstituted C₁-C₃        alkyl), -L₆-(substituted or unsubstituted C₂-C₄ alkenyl),        -L₆-(substituted or unsubstituted heteroaryl), or        -L₆-(substituted or unsubstituted aryl), wherein L₆ is a bond,        O, S, —S(═O), S(═O)₂, NH, C(O), —NHC(O)O, —OC(O)NH, —NHC(O), or        —C(O)NH;

    -   each R₉ is independently selected from among H, substituted or        unsubstituted lower alkyl, and substituted or unsubstituted        lower cycloalkyl;

    -   each R₁₀ is independently H, substituted or unsubstituted lower        alkyl, or substituted or unsubstituted lower cycloalkyl; or

    -   two R₁₀ groups can together form a 5-, 6-, 7-, or 8-membered        heterocyclic ring; or

    -   R₁₀ and R₁₁ can together form a 5-, 6-, 7-, or 8-membered        heterocyclic ring; or

    -   each R₁₁ is independently selected from H or alkyl; and        pharmaceutically active metabolites, pharmaceutically acceptable        solvates, pharmaceutically acceptable salts, or pharmaceutically        acceptable prodrugs thereof.

In some embodiments, the compound of Formula (A) is a BTK inhibitor. Insome embodiments, the compound of Formula (A) is an ITK inhibitor. Insome embodiments, the compound of Formula (A) inhibits ITK and BTK.

In some embodiments, the compound of Formula (A) has the structure:

wherein:

-   -   A is N;    -   R₂ and R₃ are each H;    -   R₁ is phenyl-O-phenyl or phenyl-S-phenyl; and    -   R₄ is L₃-X-L₄-G, wherein,    -   L₃ is optional, and when present is a bond, optionally        substituted or unsubstituted alkyl, optionally substituted or        unsubstituted cycloalkyl, optionally substituted or        unsubstituted alkenyl, optionally substituted or unsubstituted        alkynyl;    -   X is optional, and when present is a bond, O, —C(═O), S, —S(═O),        —S(═O)₂, —NH, —NR₉, —NHC(O), —C(O)NH, —NR₉C(O), —C(O)NR₉,        —S(═O)₂NH, —NHS(═O)₂, —S(═O)₂NR₉—, —NR₉S(═O)₂, —OC(O)NH—,        —NHC(O)O—, —OC(O)NR₉—, —NR₉C(O)O—, —CH═NO—, —ON═CH—,        —NR₁₀C(O)NR₁₀—, heteroaryl, aryl, —NR₁₀C(═NR₁₁)NR₁₀—,        —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or —C(═NR₁₁)O—;    -   L₄ is optional, and when present is a bond, substituted or        unsubstituted alkyl, substituted or unsubstituted cycloalkyl,        substituted or unsubstituted alkenyl, substituted or        unsubstituted alkynyl, substituted or unsubstituted aryl,        substituted or unsubstituted heteroaryl, substituted or        unsubstituted heterocycle;    -   or L₃, X and L₄ taken together form a nitrogen containing        heterocyclic ring;    -   G is

wherein,

-   -   R₆, R₇ and R₈ are independently selected from among H, lower        alkyl or substituted lower alkyl, lower heteroalkyl or        substituted lower heteroalkyl, substituted or unsubstituted        lower cycloalkyl, and substituted or unsubstituted lower        heterocycloalkyl.

In some embodiments, the ACK inhibitor is(R)-1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(i.e. PCI-32765/ibrutinib)

In some embodiments, the ACK inhibitor is AVL-263 (AvilaTherapeutics/Celgene Corporation), AVL-292 (Avila Therapeutics/CelgeneCorporation), AVL-291 (Avila Therapeutics/Celgene Corporation),BMS-488516 (Bristol-Myers Squibb), BMS-509744 (Bristol-Myers Squibb),CGI-1746 (CGI Pharma/Gilead Sciences), CTA-056, GDC-0834 (Genentech),HY-11066 (also, CTK4I7891, HMS3265G21, HMS3265G22, HMS3265H21,HMS3265H22, 439574-61-5, AG-F-54930), ONO-4059 (Ono Pharmaceutical Co.,Ltd.), ONO-WG37 (Ono Pharmaceutical Co., Ltd.), PLS-123 (PekingUniversity), RN486 (Hoffmann-La Roche), or HM71224 (Hanmi PharmaceuticalCompany Limited).

In some embodiments, the ACK inhibitor is4-(tert-butyl)-N-(2-methyl-3-(4-methyl-6-((4-(morpholine-4-carbonyl)phenyl)amino)-5-oxo-4,5-dihydropyrazin-2-yl)phenyl)benzamide(CGI-1746);7-benzyl-1-(3-(piperidin-1-yl)propyl)-2-(4-(pyridin-4-yl)phenyl)-1H-imidazo[4,5-g]quinoxalin-6(5H)-one(CTA-056);(R)—N-(3-(6-(4-(1,4-dimethyl-3-oxopiperazin-2-yl)phenylamino)-4-methyl-5-oxo-4,5-dihydropyrazin-2-yl)-2-methylphenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide(GDC-0834);6-cyclopropyl-8-fluoro-2-(2-hydroxymethyl-3-{1-methyl-5-[5-(4-methyl-piperazin-1-yl)-pyridin-2-ylamino]-6-oxo-1,6-dihydro-pyridin-3-yl}-phenyl)-2H-isoquinolin-1-one(RN-486);N-[5-[5-(4-acetylpiperazine-1-carbonyl)-4-methoxy-2-methylphenyl]sulfanyl-1,3-thiazol-2-yl]-4-[(3,3-dimethylbutan-2-ylamino)methyl]benzamide(BMS-509744, HY-11092); orN-(5-((5-(4-Acetylpiperazine-1-carbonyl)-4-methoxy-2-methylphenyl)thio)thiazol-2-yl)-4-4(3-methylbutan-2-yl)amino)methyl)benzamide(HY11066).

In some embodiments, the ACK inhibitor is:

In some embodiments, ACK inhibitor is an ITK inhibitor. In someembodiments, the ITK inhibitor is an ITK inhibitor compound described inWO2002/0500071, which is incorporated by reference in its entirety. Insome embodiments, the ITK inhibitor is an ITK inhibitor compounddescribed in WO2005/070420, which is incorporated by reference in itsentirety. In some embodiments, the ITK inhibitor is an ITK inhibitorcompound described in WO2005/079791, which is incorporated by referencein its entirety. In some embodiments, the ITK inhibitor is an ITKinhibitor compound described in WO2007/076228, which is incorporated byreference in its entirety. In some embodiments, the ITK inhibitor is anITK inhibitor compound described in WO2007/058832, which is incorporatedby reference in its entirety. In some embodiments, the ITK inhibitor isan ITK inhibitor compound described in WO2004/016610, which isincorporated by reference in its entirety. In some embodiments, the ITKinhibitor is an ITK inhibitor compound described in WO2004/016611, whichis incorporated by reference in its entirety. In some embodiments, theITK inhibitor is an ITK inhibitor compound described in WO2004/016600,which is incorporated by reference in its entirety. In some embodiments,the ITK inhibitor is an ITK inhibitor compound described inWO2004/016615, which is incorporated by reference in its entirety. Insome embodiments, the ITK inhibitor is an ITK inhibitor compounddescribed in WO2005/026175, which is incorporated by reference in itsentirety. In some embodiments, the ITK inhibitor is an ITK inhibitorcompound described in WO2006/065946, which is incorporated by referencein its entirety. In some embodiments, the ITK inhibitor is an ITKinhibitor compound described in WO2007/027594, which is incorporated byreference in its entirety. In some embodiments, the ITK inhibitor is anITK inhibitor compound described in WO2007/017455, which is incorporatedby reference in its entirety. In some embodiments, the ITK inhibitor isan ITK inhibitor compound described in WO2008/025820, which isincorporated by reference in its entirety. In some embodiments, the ITKinhibitor is an ITK inhibitor compound described in WO2008/025821, whichis incorporated by reference in its entirety. In some embodiments, theITK inhibitor is an ITK inhibitor compound described in WO2008/025822,which is incorporated by reference in its entirety. In some embodiments,the ITK inhibitor is an ITK inhibitor compound described inWO2011/017219, which is incorporated by reference in its entirety. Insome embodiments, the ITK inhibitor is an ITK inhibitor compounddescribed in WO2011/090760, which is incorporated by reference in itsentirety. In some embodiments, the ITK inhibitor is an ITK inhibitorcompound described in WO2009/158571, which is incorporated by referencein its entirety. In some embodiments, the ITK inhibitor is an ITKinhibitor compound described in WO2009/051822, which is incorporated byreference in its entirety. In some embodiments, the ITK inhibitor is anITK inhibitor compound described in U.S. Ser. No. 13/177,657, which isincorporated by reference in its entirety.

In some embodiments, the ITK inhibitor has a structure selected from thegroup consisting of:

Pharmaceutical Compositions/Formulations

Disclosed herein, in certain embodiments, are compositions comprising atherapeutically effective amount of an ACK inhibitor compound, and apharmaceutically acceptable excipient. In some embodiments, the ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as for exampleibrutinib) is a compound of Formula (A). In some embodiments, the ACKinhibitor compound is(R)-1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(i.e. PCI-32765/ibrutinib).

Pharmaceutical compositions of ACK inhibitor compound (e.g., an ITK orBTK inhibitor, such as for example ibrutinib) are formulated in aconventional manner using one or more physiologically acceptablecarriers including excipients and auxiliaries which facilitateprocessing of the active compounds into preparations which can be usedpharmaceutically. Proper formulation is dependent upon the route ofadministration chosen. A summary of pharmaceutical compositionsdescribed herein is found, for example, in Remington: The Science andPractice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack PublishingCompany, 1995); Hoover, John E., Remington's Pharmaceutical Sciences,Mack Publishing Co., Easton, Pa. 1975; Liberman, H. A. and Lachman, L.,Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980;and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Ed.(Lippincott Williams & Wilkins 1999).

A pharmaceutical composition, as used herein, refers to a mixture of anACK inhibitor compound (e.g., an ITK or BTK inhibitor, such as forexample ibrutinib) with other chemical components, such as carriers,stabilizers, diluents, dispersing agents, suspending agents, thickeningagents, and/or excipients.

Pharmaceutical compositions are optionally manufactured in aconventional manner, such as, by way of example only, by means ofconventional mixing, dissolving, granulating, dragee-making, levigating,emulsifying, encapsulating, entrapping or compression processes.

The pharmaceutical formulations described herein are administered by anysuitable administration route, including but not limited to, oral,parenteral (e.g., intravenous, subcutaneous, intramuscular), intranasal,buccal, topical, rectal, or transdermal administration routes.

The pharmaceutical compositions described herein are formulated into anysuitable dosage form, including but not limited to, aqueous oraldispersions, liquids, gels, syrups, elixirs, slurries, suspensions andthe like, for oral ingestion by an individual to be treated, solid oraldosage forms, aerosols, controlled release formulations, fast meltformulations, effervescent formulations, lyophilized formulations,tablets, powders, pills, dragees, capsules, delayed releaseformulations, extended release formulations, pulsatile releaseformulations, multiparticulate formulations, and mixed immediate releaseand controlled release formulations. In some embodiments, thecompositions are formulated into capsules. In some embodiments, thecompositions are formulated into solutions (for example, for IVadministration).

The pharmaceutical solid dosage forms described herein optionallyinclude a compound described herein and one or more pharmaceuticallyacceptable additives such as a compatible carrier, binder, fillingagent, suspending agent, flavoring agent, sweetening agent,disintegrating agent, dispersing agent, surfactant, lubricant, colorant,diluent, solubilizer, moistening agent, plasticizer, stabilizer,penetration enhancer, wetting agent, anti-foaming agent, antioxidant,preservative, or one or more combination thereof.

In some embodiments, using standard coating procedures, such as thosedescribed in Remington's Pharmaceutical Sciences, 20th Edition (2000), afilm coating is provided around the compositions. In some embodiments,the compositions are formulated into particles (for example foradministration by capsule) and some or all of the particles are coated.In some embodiments, the compositions are formulated into particles (forexample for administration by capsule) and some or all of the particlesare microencapsulated. In some embodiments, the compositions areformulated into particles (for example for administration by capsule)and some or all of the particles are not microencapsulated and areuncoated.

In some embodiments, the pharmaceutical compositions are formulated suchthat the amount of the ACK inhibitor (e.g., an ITK or BTK inhibitor,such as for example ibrutinib) in each unit dosage form is about 140 mgper unit.

Kits/Articles of Manufacture

Described herein are kits for preventing the occurrence of graft versushost disease (GVHD) or reducing the severity of GVHD occurrence in apatient requiring cell transplantation comprising atherapeutically-effective amount of an ACK inhibitor compound (e.g., anITK or BTK inhibitor, such as for example ibrutinib).

Further described herein are kits for treating a patient for alleviationof a bone marrow mediated disease, with alleviation of consequentlydeveloped graft versus host disease (GVHD) comprising a therapeuticallyeffective amount of an ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as for example ibrutinib), wherein a therapeuticallyeffective amount of an ACK inhibitor compound (e.g., an ITK or BTKinhibitor, such as for example ibrutinib) is administered prior to orconcurrently with allogeneic hematopoietic stem cells and/or allogeneicT-cells.

For use in the therapeutic applications described herein, kits andarticles of manufacture are also described herein. In some embodiments,such kits include a carrier, package, or container that iscompartmentalized to receive one or more containers such as vials,tubes, and the like, each of the container(s) including one of theseparate elements to be used in a method described herein. Suitablecontainers include, for example, bottles, vials, syringes, and testtubes. The containers can be formed from a variety of materials such asglass or plastic.

The articles of manufacture provided herein contain packaging materials.Examples of pharmaceutical packaging materials include, but are notlimited to, blister packs, bottles, tubes, inhalers, pumps, bags, vials,containers, syringes, bottles, and any packaging material suitable for aselected formulation and intended mode of administration and treatment.A wide array of formulations of the compounds and compositions providedherein are contemplated as are a variety of treatments for any disorderthat benefit by inhibition of BTK, or in which BTK is a mediator orcontributor to the symptoms or cause.

The container(s) optionally have a sterile access port (for example thecontainer is an intravenous solution bag or a vial having a stopperpierceable by a hypodermic injection needle). Such kits optionallycomprise a compound with an identifying description or label orinstructions relating to its use in the methods described herein.

A kit will typically include one or more additional containers, eachwith one or more of various materials (such as reagents, optionally inconcentrated form, and/or devices) desirable from a commercial and userstandpoint for use of a compound described herein. Non-limiting examplesof such materials include, but are not limited to, buffers, diluents,filters, needles, syringes, carrier, package, container, vial and/ortube labels listing contents and/or instructions for use, and packageinserts with instructions for use. A set of instructions will alsotypically be included.

In some embodiments, a label is on or associated with the container. Alabel can be on a container when letters, numbers or other charactersforming the label are attached, molded or etched into the containeritself; a label can be associated with a container when it is presentwithin a receptacle or carrier that also holds the container, e.g., as apackage insert. A label can be used to indicate that the contents are tobe used for a specific therapeutic application. The label can alsoindicate directions for use of the contents, such as in the methodsdescribed herein.

In certain embodiments, a pharmaceutical composition comprising the ACKinhibitor compound (e.g., an ITK or BTK inhibitor, such as for exampleibrutinib) is presented in a pack or dispenser device which can containone or more unit dosage forms. The pack can for example contain metal orplastic foil, such as a blister pack. The pack or dispenser device canbe accompanied by instructions for administration. The pack or dispensercan also be accompanied with a notice associated with the container inform prescribed by a governmental agency regulating the manufacture,use, or sale of pharmaceuticals, which notice is reflective of approvalby the agency of the form of the drug for human or veterinaryadministration. Such notice, for example, can be the labeling approvedby the U.S. Food and Drug Administration for prescription drugs, or theapproved product insert. Compositions containing a compound providedherein formulated in a compatible pharmaceutical carrier can also beprepared, placed in an appropriate container, and labeled for treatmentof an indicated condition.

EXAMPLES Example 1: Immune Reconstitution and the Development of cGVHD

The influence of lymphocyte reconstitution at days 30 and 100 followingallogeneic SCT on subsequent development of cGVHD was evaluated using ofan extensive immune reconstitution flow cytometric “immunome” assay,which allowed for monitoring of changes in cell activation markers,memory T cell status, Treg subsets, NK cell subsets and Th1 vs Th2 cellsubsets. Patients who developed cGVHD had a larger increase in CD4+ Tcells and a smaller increase in CD8+ T cells compared with patients whodid not develop cGVHD over time, suggesting a selective expansion ofCD4+ T cells. Further, a significant decrease in NK cells andconcomitant increase in percentages of activated B cells was noted. Anincrease in CD4+ cells is associated with an inflammatory phenotype, anda Th2-skewed proinflammatory response may contribute to B cellactivation. The presence of a Th2-skewed phenotype was supported by thepresence of increased CD4+/CD193+ cells among patients with cGVHD, asCCR3 is preferentially expressed on Th2 cells.

Example 2: Ibrutinib in a Murine Model of cGVHD

An established in vivo allo-bone marrow transplant (BMT) model systemwas used to preclinically test ibrutinib as a therapy for cGVHD. TheLP/J→C57BL/6 model is a murine model of sclerodermatous cGVHD whichdevelops dermal lesions characterized by hair loss, redness, flaking,scabbing, hunched posture, and thickened skin. In this murine model,external symptoms become apparent between days 20 and 25 and peakbetween days 37 and 47 post hematopoietic stem cell transplantation(HSCT).

C57BL/6 mice received lethal X-ray irradiation (850cGy) followed byallo-BMT derived from MHC-matched LP/J mice. A small number of maturespleen cells were included in the transplant to seed the development ofcGVHD. The studies demonstrated that approximately 1/3 of mice survivedto day 25 post transplant and at that time began to develop classicexternal signs of cGVHD including scleroderma, hair loss, hunchedposture, weight loss, and dermal fibrosis. Therefore, 25 days post-BMTwas selected for treatment time point.

Ibrutinib Ameliorates cGVHD Symptomatology after Allotransplant

C57BL/6 mice were engrafted with LP/J bone marrow after 850cGy lethalirradiation. 25 days post-transplant mice were randomly assigned tovehicle, cyclosporine, or ibrutinib cohorts and drug was administeredvia drinking water or intraperitoneal injection. Scoring was conductedon day 36 or day 39 post-transplantation using a physical scoring systemadapted from Cooke et al., which incorporates weight, posture, coatcondition, skin condition, and mobility.

These studies confirmed a dramatic therapeutic response to ibrutinibwhich allowed for complete resolution of cGVHD induced scleroderma,alopecia, weight loss, and paralysis as compared to vehicle orcyclosporine treatment groups (FIGS. 1A, D). The quality of therapeuticresponse was quantitatively assessed using a previously establishedscoring model for the assessment of hair loss, scleroderma, weight loss,posture, and mobility in mice suffering from cGVHD (FIG. 1B). In thescoring model, scores range from 0 (healthy mouse) to 19 (mouse whichhas died due to cGVHD) with 18 representing the maximum score for aliving mouse with cGVHD (FIG. 1C). cGVHD progression was defined as a >2point change in overall cGVHD score from treatment baseline. Histologicpreparations of sclerodermatous skin lesions revealed dermal fibrosis,epidermal hyperplasia, serocellular crusting, erosion, andlymphohistiocytic infiltration, consistent with external examination(FIG. 1E). Normal dermal histology was observed in mice receivingtherapeutic ibrutinib.

Ibrutinib significantly extended median time to cGVHD progression by 14days and 33% (6 of 18) of ibrutinib treated mice remained progressionfree as compared to 12% (2 of 18) of mice receiving vehicle and 10% (1of 11) of mice receiving cyclosporine 10 mg/kg/day (p<0.02) (FIG. 5). A100% survival in the ibrutinib cohort as compared to 82 and 88% survivalfor cyclosporine and vehicle groups respectively, was observed. Weeklyevaluation of mouse bodyweight revealed little variation between groupswith ibrutinib treated mice weighing slightly more on average.

Tregs were not Inhibited by Ibrutinib

In allo-BMT recipients, Tregs (regulatory T cells) control cGVHD byactively suppressing autoreactive T-cells within the periphery;unfortunately, most current therapies disrupt Treg development orfunctionality. To study the effects of ibrutinib on Tregs, C57BL/6 micewere treated with ibrutinib (25 mg/kg/day) or vehicle for 9 weeks andthe percent FoxP3+ CD4+ cells was analyzed by flow cytometry onperipheral blood. In addition, purified CD4+ CD25hiCD127dim CD49d-FoxP3+Tregs were pretreated with 1 μM ibrutinib or vehicle and mixed with CFSE-labeled autologous CD8+ responder cells at different responder:suppressor ratios of 1:0, 1:1, 2:1, 4:1, 8:1, and 16:1.Anti-CD3/CD28/CD2 stimulation beads were added and stimulation wasassessed by CFSE (carboxyfluorescein succinimidyl ester) dilutioncalculated division index after 6 days. Negative control wells containedno stimulation beads.

The in vivo data showed that ibrutinib did not diminish overall Tregnumbers after 9 weeks of continuous ibrutinib therapy (FIG. 2A). The invitro data indicated that the suppressive function of human Tregs wasmaintained after ibrutinib treatment as assayed by an in vitro T-cellsuppression assay (FIG. 2B). The data showed that ibrutinib had theability to suppress anti-host immunity while preserving Treg function,which is important for the graft-versus-tumor effect.

Th2 Immunity was Inhibited by Ibrutinib

Intracellular staining was performed for IFNγ and IL4 inibrutinib-treated, TCR-stimulated CD4+ T-cells. Following stimulation, asignificant decrease was identified in the IL4-producing Th2 populationof CD4+ T-cells, whereas IFNγ-producing Th1 cells were largelyunaffected (FIG. 3A). These data confirmed that a significant divergenceof the two cell populations was achieved in a purified T-cell culture atibrutinib doses ranging from 0.1-1 μM. This dose range was consistentwith serum concentrations observed in vivo during pharmacokineticstudies of ibrutinib in both mouse and human trials. To assess thelong-term implications of ibrutinib-induced Th1 cytokine skewing, IgGsubisotype analyses were conducted in a cohort of 8 month old C57BL/6EμCL1 mice. These mice were treated continuously for 7 months withibrutinib (25 mg/kg/day) or vehicle. Results revealed a significant(p=0.002) inversion of the Th1/Th2 ratio as measured by the relativelevels of IgG1 (Th2) and IgG2c (Th1), confirming an in vivoibrutinib-related Th1 skewing (FIG. 3B).

Th17 Immunity was Inhibited by Ibrutinib

In cGVHD, there is a common link in which alloreactive Th2 and Th17T-cells drive pro-fibrotic pathways and B-cell autoantibody productionas a result of defective thymic conditioning. Given the role of Th17cells, the effect of ibrutinib on this specific T-cell subtype wasinvestigated. Healthy donor Th17 cells were magnetically isolated fromfreshly isolated healthy donor PBMCs using CXCR3-CD4+CCL6+ isolation andTCR stimulation for 12 hours following 30 minute pretreatment withvehicle or 1 μM ibrutinib. The percentage of IL17 secreting CD4+ T-cellswas quantified by intracellular cytokine staining and normalized tovehicle treatment (FIG. 4). The data indicated that ibrutinib limitedthe TCR-induced activation of Th17 cells.

Ibrutinib Therapeutically Controlled cGVHD-Induced Organ Injury

In addition to the externally measurable cGVHD metrics, it was foundthat the LP/J→C57BL/6 model developed pulmonary and renal cGVHD,apparent upon histologic assessment. Evaluation of H&E stained sectionsrevealed that ibrutinib therapy systemically limited cGVHD-inducedaggregates of lymphocytes, plasma cells, and histiocytes surroundingbronchioles and small caliber vessels throughout the pulmonaryparenchyma and within the renal interstitium. Immunohistochemistryrevealed B220+ B-cell and CD3+ T-cell pulmonary infiltration in additionto CD3+ T-cell renal infiltration in both the vehicle and cyclosporinegroups which was not observed in ibrutinib treatment groups (FIG. 6A).Coded pathologic analysis by a trained veterinary pathologist confirmedthat ibrutinib improved internal systemic cGVHD in this model (FIGS. 6Band C).

An additional long-term therapeutic experiment was conducted (FIG. 6D).Once again, ibrutinib significantly limited cGVHD progression ascompared with vehicle control (P=0.0019). It was also found thatwithdrawal of therapy at day 60 permitted clinical breakthrough cGVHD ina single mouse (1 of 6); however, this was not statisticallysignificant. A similar trend was observed by external cGVHD scoring.Analysis of internal cGVHD pathology within the pulmonary and renaltissues on day 75 suggested that continuous long-term ibrutinib was moreeffective at controlling cGVHD; notably, internal pathology of the lungand kidney was not curtailed in BM-only recipients, indicating thatcertain cGVHD internal pathology in this model persists despite theelimination of T cells from the graft similar to what is observed inhuman allo-HSCT recipients. Prophylactic ibrutinib treatment initiatedpre-HSCT at day −2 and concluded at day 25 did not yield a significantimprovement in cGVHD progression, suggesting that ibrutinib will be mosteffective when T and B cell responses are more fully developed.

Example 3: Ibrutinib Inhibition of CD4 T-Cell and B-Cell Activation inCells from Patients with cGVHD

CD4+ T-Cell Activation in Cells from cGVHD Patients Inhibited byIbrutinib

Primary CD4+ T-cells were isolated from patients with active cGVHD,pretreated with 1 μM ibrutinib (or DMSO), and stimulated using anti-CD3for 6 hours. Quantitation of the activated T-cell percentage for eachpatient showed a significant decrease in the population of CD69+ CD4+T-cells (FIG. 7A) in cells pretreated with ibrutinib as compared toDMSO.

B-Cell Activation in Cells from cGVHD Patients Inhibited by Ibrutinib

B-cells isolated from patients with cGVHD were pretreated with 1 μMibrutinib and stimulated with anti-IgM for 45 minutes. Analysis ofB-cell receptor pathway activation using phospho-specific antibodies forpERK1/2, pBTK, and pPLCγ2 revealed that ibrutinib was effective ininhibiting the B-cell receptor pathway (FIG. 7B). These data confirmedthat ibrutinib curtailed immune receptor activation in human B and Tcells in the setting of active cGVHD.

Example 4: Clinical Studies of Ibrutinib in CLL and Lymphoma

Fifty-six patients, including 16 with CLL, were treated on the initialphase I study of ibrutinib. Seven cohorts were evaluated: 5 cohortswhere patients were treated on a 28 days on, 7 days off schedule, and 2cohorts where patients were treated on a continuous dosing schedule at8.3 mg/kg daily or a fixed dose of 560 mg once daily. Patients hadreceived a median of 3 prior regimens, and the median age was 65, withthe oldest patient being 82 years old. The maximum tolerated dose wasnot reached, and only 2 dose-limiting toxicities were observed: a grade3 allergic hypersensitivity in a patient with a history of drughypersensitivities; and a dose interruption for more than 7 days becauseof transient grade 2 neutropenia. Using a competitive binding assay toevaluate, ≥95% BTK occupancy was achieved 4 hours post dose in allpatients receiving 2.5 mg/kg/day. Thus, doses of 420 mg and 840 mg givendaily were selected for further study. Of 50 patients evaluable fortumor response, 60% achieved an objective response (CR or PR). Responseswere observed across all histologies, including in 11 of the 16 patientswith CLL/SLL. All of the patients with CLL who responded had rapidreduction in lymphadenopathy during the first cycle accompanied by anincrease in the absolute lymphocyte count, and all but one had aneventual reduction in the ALC to meet IWCLL criteria for a PR. Responseswere durable, with a median progression-free survival of 13.6 months.

Based on the impressive responses in CLL patients seen in the phase Istudy, a phase Ib/II study was conducted in patients with CLL. Patientswere enrolled in one of 5 cohorts evaluating ibrutinib at a fixed doseof 420 mg daily or 840 mg daily. Cohorts evaluated patients who weretreatment-naïve and at or above the age of 65, relapsed or refractoryafter 2 or more prior lines of treatment, including a purine-nucleosideanalog, or high-risk, with relapse within 2 years of receivingchemoimmunotherapy, or the presence of del17p. One-hundred-and-sixteenpatients were enrolled: 31 treatment-naïve patients, 61 in therelapsed/refractory cohorts, and 24 high-risk patients. The overallmedian follow-up was 16.6 months, with 4 median prior therapies in boththe relapsed/refractory and high-risk cohorts. The most common adverseevents noted were diarrhea, fatigue, upper respiratory tract infection,rash, nausea, and arthralgia, and most were grade 2 or less.Importantly, no evidence of cumulative toxicity has been reported.Responses were observed, independent of poor-risk factors includingadvanced disease stage, increasing numbers of prior therapies, higherbeta-2-microglobulin, or poor-risk cytogenetics, with an ORR of 67% inpatients with del17p in the relapsed/refractory cohorts. The estimated22 month PFS for the 85 patients in the relapsed/refractory andhigh-risk cohorts was 76% and was 96% for the 31 treatment-naïvepatients. The estimated 22 month overall survival for these two groupsrespectively was 85% and 96%. Median progression-free and overallsurvivals for any of the cohorts had not been met at the time.

Example 5: Clinical Study of Ibrutinib in Patients with SteroidResistant or Refractory Chronic Graft-Versus-Host Disease (cGVHD)

cGVHD and its associated immune deficiency has been identified as aleading cause of non-relapse mortality (NRM) in allogeneic SCTsurvivors. SCT survivors with cGVHD are 4.7 times as likely to developsevere or life-threatening health conditions compared with healthysiblings, and patients with active cGVHD are more likely to reportadverse general health, mental health, functional impairments, activitylimitation, and pain than allo-SCT survivors with no history of cGVHD.Rituximab notwithstanding, historical response rates with a number ofinvestigational agents in steroid-refractory cGVHD have been around 30%,so this is a patient population with a clear need for an effectiveintervention to reduce dependence on steroids and improve quality oflife and survival. Ibrutinib induces apoptosis in B lymphocytes throughinhibition of the BCR pathway and antagonizes multiple externalmicroenvironment survival signals mediated through cytokines such asBAFF, and it can reverse Th2 polarization. It has thus far been shown tobe safe in the treatment of patients with relapsed or refractory B celllymphomas and CLL, with the most common toxicities being diarrhea,fatigue, upper respiratory infection and rash, and being grade 2 orless. Importantly, no cumulative toxicities were noted, allowinglong-term use of the drug. In comparison, long-term use of the steroidsand calcineurin inhibitors used to treat cGVHD is known to cause adverseeffects, leading to much of the morbidity and mortality seen in patientswith cGVHD. Based on ibrutinib's tolerability in early phase studies andits mechanism of action, it was expected that this would be awell-tolerated agent and with clinical efficacy against cGVHD.

For the primary objectives of evaluating safety and efficacy ofibrutinib when used for cGVHD, it is expected that ibrutinib will bewell tolerated in patients with steroid-dependent/refractory cGVHD andwill improve response at 12 weeks compared with the historical responserate of 30%. Further, it is expected that use of ibrutinib will allowfaster tapering of steroids and will contribute to improved quality oflife at 1 and 2 years compared with historical controls. Becausepatients will be exposed to less corticosteroids, it is expected thatrelapse rates of primary disease will improve with ibrutinib,particularly in patients who receive a transplant for a lymphoidmalignancy, in which ibrutinib has documented efficacy in phase IIstudies. Because ibrutinib has effects on ITK it is expected thatcompared to institutional controls, use of ibrutinib in this settingwill skew towards a Th1 phenotype, which will be evaluated throughserial evaluation of immune reconstitution by flow cytometry.Additionally, it inhibits activation of Th17 cells, while preserving thefunction and numbers of Tregs, thereby preserving the graft-versus-tumoreffect while treating GVHD.

Objectives of the study:

1. Primary endpoint:

To determine the safety of ibrutinib when given for chronic GVHD (PhaseIb portion)

To evaluate the response rate (CR+PR) at 12 weeks using ibrutinib as atreatment for steroid-refractory or -resistant chronic GVHD (Phase IIportion)

2. Secondary endpoints:

To evaluate the impact of ibrutinib on steroid dose at 12 weeks, 6months, 1 year, and 2 years

To evaluate response at 6 months and 1 and 2 years

To evaluate overall survival at 1 and 2 years

To evaluate the relapse rate (both primary disease and cGVHD symptoms)at 1 year

To evaluate the incidence of grade 3-5 infections during treatment

To evaluate quality of life at 1 and 2 years

To evaluate effect on immune reconstitution at 1 and 2 years

Eligibility Criteria:

1. Classic or overlap chronic GVHD that is resistant or refractory tocorticosteroids (equivalent to at least 0.5 mg/kg/day or 1 mg/kg/everyother day prednisone for at least one month of treatment).Organ-specific topical therapy permitted

2. History of allogeneic stem cell transplant for hematologic malignancy

3. Age 18-75 years at the time of registration

4. Within seven days of administration of the first dose of ibrutinib,the patient must have adequate organ function and performance status asfollows:

Absolute neutrophil count (ANC) ≥500/μL

Platelets ≥30,000/μL

Total bilirubin ≤2.5× institutional upper limit of normal unless due toGilbert's disease unless attributable to cGVHD

AST (SGOT)≤2.5× institutional upper limit of normal unless attributableto cGVHD

Creatinine clearance >40 mL/min

5. ECOG performance status <2

6. Life expectancy >12 weeks

7. Willing and able to participate in all required evaluations andprocedures in this study protocol

8. Able to understand the purpose and risks of the study and providesigned and dated informed consent and authorization to use protectedhealth information (in accordance with national and local subjectprivacy regulations)

Exclusion Criteria:

1. New immunosuppression within 4 weeks of starting ibrutinib

2. “Currently active” malignancy, except for adequately treated basalcell or squamous cell skin cancer, in situ cervical cancer, or othercancer from which the subject has been disease free for at least 2years, aside from the primary indication for transplant, or which willnot limit survival to less than 2 years

3. A life-threatening illness, medical condition or organ systemdysfunction which, in the investigator's opinion, could compromise thesubject's safety or put the study outcomes at undue risk

4. Active and uncontrolled bacterial, fungal, or viral infection

5. Significant cardiovascular disease such as uncontrolled orsymptomatic arrhythmias, congestive heart failure, or myocardialinfarction in absence of significant predisposing cause (i.e., severeautoimmune hemolytic anemia or sepsis) within 6 months of screening, orany Class 3 or 4 cardiac disease as defined by the New York HeartAssociation Functional Classification

6. Known history of Human Immunodeficiency Virus (HIV) or activeinfection with Hepatitis C Virus (HCV) or Hepatitis B Virus (HBV) or anyuncontrolled active systemic infection.

7. Concurrent antineoplastic therapy after hematopoietic stem celltransplant

8. Lactating or pregnant

9. Will not agree to use highly effective contraception (e.g., condoms,implants, injectables, combined oral contraceptives, some intrauterinedevices [IUDs], sexual abstinence, or sterilized partner) during thestudy and for 30 days after the last dose of study drug (Note: appliesto men and women of child-bearing potential only)

Study Design:

1. Overview

This is a non-randomized, open-label phase Ib/II study for patients withsteroid resistant or refractory chronic graft-versus-host disease(cGVHD), equivalent to at least 0.5 mg/kg/day or 1 mg/kg/every other dayprednisone for at least one month of treatment, following allogeneicstem cell transplant for hematologic malignancy. If a patient is on acalcineurin inhibitor and the level is less than 5 ng/ml at the start ofthe study, it will be stopped. Following enrollment, patients will bestarted on 420 mg ibrutinib daily, based on phase I and II studies inhematologic malignancies demonstrating that this dose is well toleratedand that 90% of the BTK active sites are occupied at this dose. For easeof documentation and follow-up, a cycle will be defined as 28 days.Treatment is oral, and will be administered on an outpatient basis. Thefirst six patients will be subject to a dose-limiting toxicity (DLT)assessment period. The DLT period will be 28 days following initialadministration of the drug, and the sixth patient must complete the DLTperiod before accrual can continue. A DLT is defined as the following:grade 2 acute graft-versus-host disease (biopsy-proven preferable butnot necessary); grade 4 thrombocytopenia that does not improve to 80% ofbaseline or better following a 14-day treatment-free period withoutdisease progression; grade 4 febrile neutropenia or infection; grade 3febrile neutropenia or infection that fails to resolve within 7 days;any Grade 4 non-hematologic toxicity excluding infection; and grade 4electrolyte abnormalities if not corrected by optimal replacementtherapy.

The initial steroid taper may start at 4 weeks following initiation ofibrutinib, but the steroid dose may not be lower than 50% of thestarting dose by the end of the third cycle (12 weeks). Patients willhave a physical exam with comprehensive cGVHD evaluation at thebeginning of each cycle. Patients will be assessed for response at theend of cycle 3, and if there is no improvement in symptoms, then thiswill be considered a treatment failure and they will come off study.Additionally, patients who require additional treatment for cGVHD priorto the response assessment at the end of cycle 3 will be considered atreatment failure, and these patients will be removed from study. Ifpatients are in a CR or PR at the 12 week assessment, they will continueon daily ibrutinib while steroids are tapered. Once the steroids havebeen tapered off, ibrutinib can be discontinued. If patients arederiving clinical benefit allowing for reduction of steroid dose at the1 year assessment point, then they will be allowed to continue on studyfor a maximum of 24 months. GVHD will be assessed monthly, andcorrelative studies, including immune reconstitution, B and T cellactivation, serum immunoglobulins, and serum BAFF levels will beassessed every 3 months. Symptom burden and quality of life studiesincluding the Lee cGVHD symptom scale, the 10-point cGVHD activityassessment, the FACT-BMT, SF36, and Human Activity Profile will beassessed at 12 weeks, 6 months, 1 year, and 2 years.

2. Ibrutinib Therapy

Ibrutinib will be administered daily each day of a 28 day cycle. Thefirst administration of ibrutinib will define C₁D1. A fixed dose of 420mg will be administered. A comprehensive chronic graft-versus-hostassessment according to NIH consensus criteria 35 will be performed atbaseline in order to determine organ-specific and global score. Thecomprehensive assessment will be repeated at the end of cycle 3. Thisstudy will be performed using a phase Ib/II study design, in which thephase II portion will be conducted as a Simon optimal 2 stage design. Ifno more than 5 of the initial 15 patients, including the 6 enrolled inthe phase Ib portion, have evidence of a CR or a PR at the 12 weekassessment, the study will be stopped for futility. At the treatingphysician's discretion, the initial taper of prednisone may begin 2weeks following initiation of ibrutinib if a clinical response is seen.The prednisone may not be tapered below 50% of the original dose by the12 week assessment period, and an increase in cGVHD symptoms duringsteroid taper requiring an increase in steroid dose to NO MORE than theoriginal dose will not be considered progression. However, if a patientrequires a steroid dose higher than the original dose or the addition ofa new treatment for cGVHD at any point, this will be considered evidenceof progressive disease and will require removal from study. Becauseapproximately 66% of patients with cGVHD are expected to progress,regardless of therapy, stopping rules will be triggered if more than 75%of patients progress prior to the assessment at the end of cycle 3.Daily ibrutinib will continue until steroids have been tapered off. Aspecific prednisone taper schedule will not be mandated. Once steroidsare off, ibrutinib will continue for an additional 4 weeks, then stop.Patients may continue on ibrutinib for 2 years, and patients who areable to stop ibrutinib will be followed for 2 years from the start oftreatment for secondary endpoints.

Endpoints/Statistical Considerations: This study will be conducted in 2parts, a phase Ib portion and a phase II portion. Six patients willinitially be enrolled to the phase Ib portion of the study. Analogous toa maximum tolerated dose (MTD) evaluation, the regimen will beconsidered sufficiently tolerable if at most one of these 6 patientsexperiences a DLT during the observation period of 28 days, in whichcase the study will progress to the phase II portion. Using a Simonoptimal two-stage phase II design, in order to test the null hypothesisthat the overall response rate will be 30% against the alternatehypothesis that the overall response rate will be 50%, with a one-sidedtype I error of 0.1 and 80% power, 32 patients will be needed. Of over100 allogeneic transplants perform annually by the inventors,approximately half of these ultimately develop cGVHD. Approximately 50%of these will have disease that is not responsive to initial treatmentwith steroids, resulting in approximately 20 incident cases of cGVHD atOhio State annually. The study is expected to accrue approximately 1patient per month, resulting in a nearly 3 year accrual period ifperformed at a single institution. Transplant volumes are slightly lowerat the University of North Carolina and the University of Chicago, andtherefore, with the addition of these 2 sites, it would be expected tobe able to fully accrue in approximately 24 months. If at most 5 of 15patients in the first stage respond at the 12 week assessment point, thestudy will be terminated. If at most 12 patients respond overall, thistreatment will not be considered worthy of further pursuit. In general,regardless of treatment, it is expected that approximately 66% ofpatients will have progressive disease by 6 months after startingtreatment. Therefore, stopping rules will be triggered if 75% ofpatients or more enrolled prior to the interim analysis cutoff developworsening cGVHD requiring treatment escalation within the first 12 weeks(3 of 4 patients, 6 of 8 patients, and 9 of 12 patients).

Response will evaluated by NIH consensus criteria for GVHD grading.

Complete response (CR) will be defined as complete resolution ofsymptoms attributable to GVHD.

Partial response (PR) will be defined as the presence of an objectiveresponse in one involved organ with no evidence of progression elsewhereand no requirements for additional systemic therapy.

The length of follow-up will be 24 months, and the estimated accrualperiod will be 2 years.

Patient characteristics will be presented as median and range forcontinuous variables and as frequency and percentage for categoricalvariables. Laboratory correlates will be summarized at each time pointusing descriptive statistics. The non-parametric Wilcoxon signed-rankprocedure will be used to compare to baseline values for correlativestudies. For the quality-of-life correlates, a 0.5 standard deviationchange will be considered statistically significant. Logistic regressionmodels will be fitted to find correlations that merit further researchin future studies. Time-course plots will be generated for each patientand repeated measures analysis of variance will be used to explorerelationships. The κ statistic will be used to evaluate agreementbetween the NIH response and clinically meaningful improvement in thequality of life measures.

Example 6: CLL/GVHD Case Study

A 52 year old male with high risk 17p del CLL was originally diagnosedin November 2002. In 2003, he was initially treated with six cycles offludarabine, cyclophosphamide, and rituximab (FCR) and achieved acomplete response. He recurred one year later with a right pleuraleffusion and retroperitoneal/mesenteric lymph nodes. In December 2006,peripheral blood fluorescence in situ hybridization (FISH) showed 23percent of the cells to be 17p deleted and CT scan showed increasingadenopathy. In March 2007, his bone marrow was hypercellular anddiffusely infiltrated with CLL (62 percent of CD45+ cells). By April2007, he was restarted on FCR and received four cycles with a partialresponse (bone marrow showing persistent disease with 44 percent CD45+cells). Because of his persistent disease and p17 del diagnosis, hereceived Campath (alemtuzumab) for a total of 20 doses and a subsequentbone marrow biopsy in October 2007 showed no CLL and the PET/CT wasnegative.

In November 2007, the patient underwent a non-myeloablative allogeneichematopoietic stem cell transplant using total lymphoid irradiation andantithymocyte globulin (TLFATG) with infusion of GCSF mobilizedperipheral blood stem cells from his matched related sibling donor(sister). His oral graft versus host disease (GVHD) prophylaxisconsisted of cyclosporine (CSA) and mycophenolate mofetil (MMF). He wastransplanted on a Stanford research protocol (BMT 172) that incorporatedrituximab 375 mg/m2 infused post-transplant on days 56, 63, 70, and 77.

The patient's post-transplantation course was complicated by infectiouscomplications (multi-lobar fungal pneumonia, influenza A, varicellazoster reactivation) and post-transplant lymphoproliferative disorder(PTLD) treated with nine doses of rituximab in 2008. The patient neverachieved full donor chimerism post-HCT. By September 2008, approximatelynine months post-HCT, he was found to have disease progression by flowcytometry and a CT scan that showed some retroperitoneal adenopathyenlargement from prior study. Due to the patient's mixed donor chimerismand disease progression, the patient received a total of five donorlymphocyte infusions (DLIs).

The patient's first DLI was given in September 2008 at a dose of 1×10⁷donor CD3+ cells/kg recipient body weight. No GVHD or disease response.He received a second DLI in November 2008 at a dose of 3×10⁷ donor CD3+cells/kg recipient body weight. He was found to have a decrease in hisallele-specific oligonucleotide (ASO) quantification results withreduction from 268,000 clonal IgH sequences per mcg of DNA down to 120.There was also an increase in donor blood T cell chimerism to 90% inJanuary 2009 with associated mild oropharyngeal chronic GVHD. Bonemarrow biopsy in April 2009 showed only 10% CLL. He received a third DLIin May 2009 at a dose of 5×10⁷ donor CD3+ cells/kg recipient bodyweight. The patient also received a cycle of rituximab (4 doses duringthis time). Approximately 18 days after his third DLI, he developed oralGVHD with erythema and ulceration associated with a dramatic increase indonor T cell chimerism from 71 percent on day of DLI infusion to 87percent 11 days later and up to 97 percent 25 days later. He did notrequire systemic steroid therapy for the GVHD but required local therapyand it persisted for about a year.

In the summer of 2009, the patient's PET/CT scan showed diseaseprogression with bulky disease in the chest/abdomen and a bone marrowbiopsy with 40 to 50 percent CLL. For this persistent disease, thepatient was subsequently treated with combination chemotherapy with fourcycles of OFAR (oxaliplatin, fludarabine, cytarabine, rituximab). InDecember 2009, the patient received a fourth DLI at a dose of 5×10⁷donor CD3+ cells/kg recipient body weight. GVHD flared when his donorchimerism reached above 95 percent following OFAR. However, his diseasepersisted, so he was given a fifth DLI in February 2010 at a dose of1×10⁶ donor CD3+ cells/kg recipient body weight. His oral GVHD requiredlocal steroid treatment. His bone marrow biopsy performed in May stillshowed 50 percent CLL.

By August 2010, the patient's PET/CT showed rapidly progressive diseaseand recurrence of massive lymphadenopathy including an abdominal mass upto 12 centimeters in diameter. In September 2010, the patient wasenrolled in a clinical trial with ibrutinib (oral BTK inhibitor) throughStanford's Hematology group and completed over three years of therapy onibrutinib, and achieved a complete response in both the bone marrow andon CT (see FIG. 8). In addition, his oral GVHD symptoms completelyresolved and he achieved a durable full donor chimerism (see FIG. 8).

In summary, this post-allogeneic HCT CLL patient had refractory CLL withoropharyngeal chronic GVHD that resolved with ibrutinib therapy. His CLLwas undetectable using B cell IgH sequencing (CLONOSIGHT minimalresidual disease test (Sequenta, Inc.)) and he achieved full donorengraftment with no chronic GVHD.

Example 7: Ibrutinib Treatment of Relapsed CLL Following AllogeneicTransplantation: Sustained Disease Response and Promising Donor ImmuneModulation

This example demonstrates the effects of ibrutinib salvage therapy in 5CLL patients who relapsed following allogeneic hematopoietic celltransplantation (allo-HCT). In addition to minimal residual disease(MRD) response measurements, donor T cell chimerism and donor B cellimmune reconstitution following ibrutinib therapy were also assessed.Five patients with high-risk CLL relapsed 1-8.5 yrs following allo-HCT.Four patients had never achieved donor CD3 T cell chimerism >95%following reduced-intensity transplant. Ibrutinib 420 mg daily wasstarted 1 mo-2 yrs after clinical relapse. Four of the 5 patientsremained on ibrutinib with treatment courses ranging from 3-17 mos. CLLMRD was measured by IgH high-throughput sequencing (HTS) using theCLONOSIGHT minimal residual disease test (Sequenta, Inc.), which has thesensitivity to detect 1 CLL clone per million leukocytes. Lymph node(LN) size was assessed by CT scan and reported as the sum of theproducts of the LN diameters (SPD). Donor B cell reconstitution wasdetermined by IgH HTS quantification of total IgH molecules and uniqueIgH clonotypes. Lymphocytosis was observed in all 5 patients followinginitiation of ibrutinib treatment, consistent with previous reports. In2 patients who received >1 yr of ibrutinib treatment, lymphocytosispeaked at 3 wks and 8 wks after initiation of treatment and slowlydeclined thereafter, fully resolving within 1 yr (FIG. 9A). All 4patients with pathologic lymphadenopathy prior to treatment experienceddramatic LN reduction (FIG. 9B; 68% average reduction in LN size after 3months on ibrutinib). The longest duration of follow-up was reported forpatient SPN 3975, who had a 17p deletion and received ibrutinib for 39months. Treatment was discontinued after CLL MRD became undetectable byCLONOSIGHT minimal residual disease test (FIG. 9C). Evidence of donor Tcell immune modulation included achievement of full donor CD3 chimerismafter 1 year and resolution of oral and skin chronic graft-versus-hostdisease (GVHD) after 6 months. Although this patient has been offibrutinib for >8 months, full donor chimerism persisted and CLL MRDremained undetectable (FIG. 9C). Before ibrutinib treatment, donor Bcells (excluding the patient's CLL clone) accounted for <0.2% of totalPBMC as determined by IgH HTS. Following discontinuation of ibrutinib,the percentage of donor B cells increased within 6 months to >1% of PBMC(FIG. 9D). Furthermore, recovering B cells had diverse, low frequencyIgH clonotypes (FIG. 9E). Together, these findings show rapid,sustained, and diverse immune reconstitution without CLL recurrencefollowing ibrutinib discontinuation. Ibrutinib provided effectivesalvage therapy for CLL relapse following allo-HCT. Post-transplant CLLrelapse is often extra-nodal and our experience shows ibrutinib iseffective in clearing both nodal and extra-nodal disease. One patientwho stopped therapy after achieving MRD negativity maintainsundetectable disease 8 months following ibrutinib discontinuation.Ibrutinib treatment demonstrated promising donor immune modulation bypromoting full donor chimerism and resolution of chronic GVHD. Thesedata supported using ibrutinib in relapsed CLL patients followingallo-HCT.

Example 8: Safety and Efficacy of Ibrutinib in Patients withRelapsed/Refractory (R/R) Chronic Lymphocytic Leukemia (CLL)/SmallLymphocytic Lymphoma (SLL) Who have Undergone Prior Allogeneic Stem CellTransplant

Patients with CLL who relapse after allogeneic hematopoietic stem celltransplantation (alloHCT) are difficult to treat with chemotherapy dueto impaired hematopoietic reserve, infections, and concern forgraft-versus-host disease (GVHD). Ibrutinib is approved in the USA forpatients with CLL or MCL who have received >1 prior therapy, and forpatients with CLL with del17p. In preclinical studies, ibrutinibreversed established chronic GVHD (cGVHD). The safety and efficacy ofibrutinib in a subset of patients with prior alloHCT were evaluated inthis example. Data were collected for R/R patients with prior allogeneicHSCT enrolled in 1 of 4 clinical trials (PCYC-1102, PCYC-1109,PCYC-1112, and PCYC-1117). PCYC-1112 and PCYC-1117 only enrolledpatients >6 months post-HCT and without GVHD. Efficacy evaluationsincluded overall response rate (ORR; iwCLL criteria), duration ofresponse (DOR), progression-free survival (PFS), and overall survival(OS). Safety evaluations included adverse events (AEs), includingserious AEs (SAEs). 16 patients from 4 clinical trials had prior alloHCT(median age, 54.5 y; 16 patients with ECOG performance status 0 or 1; 10patients with del17p, 3 patients with del11q, 12 patients with ≥4 priortherapies). Median time since the most recent HCT was 27 months (range,8-115). Baseline neutropenia, anemia, and thrombocytopenia were reportedin 31%, 25%, and 38%, respectively. Median time on ibrutinib was 18.1months (range, 0.4-38.8), with 12 patients being treated for >12 months.At data cut-off, 11 patients were continuing treatment. Reasons fordiscontinuation included disease progression (n=2), AEs (n=2), andconsent withdrawal (n=1). Investigator-assessed responses included 2complete responses, 9 partial responses (PRs), and 3 PRs withlymphocytosis, resulting in a best ORR of 87.5%. Median DOR, PFS, and OSwere not reached at a median follow-up of 23 months. The 24-month PFSand OS rates were 77% and 75%, respectively. Treatment-emergent grade >3SAEs were observed in 11 patients and included infections (n=6), andfebrile neutropenia, atrial flutter, colitis, perirenal hematoma,subdural hematoma, postprocedural hemorrhage, hypercalcemia, bonelesion, syncope, hematuria, urinary retention, and dyspnea (n=1 each,some events reported for the same patient). The only AE leading toibrutinib discontinuation was pneumonia (n=2); both were fatal events.Two additional deaths occurred on study due to disease progression at 24and 28 months. Ibrutinib was well tolerated in patients who had prioralloHCT, with a safety profile similar to that observed in the overallR/R CLL population. Best ORR (87.5%) was consistent with resultsobserved in the overall/broader population.

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention. It is intended thatthe following claims define the scope of the invention and that methodsand structures within the scope of these claims and their equivalents becovered thereby.

1. Use of a compound of Formula (A) for preventing the occurrence ofgraft versus host disease (GVHD) or reducing the severity of GVHDoccurrence in a patient requiring cell transplantation, wherein Formula(A) has the structure:

wherein: A is N; R₁ is phenyl-O-phenyl or phenyl-S-phenyl; R₂ and R₃ areindependently H; R₄ is L₃-X-L₄-G, wherein, L₃ is optional, and whenpresent is a bond, optionally substituted or unsubstituted alkyl,optionally substituted or unsubstituted cycloalkyl, optionallysubstituted or unsubstituted alkenyl, optionally substituted orunsubstituted alkynyl; X is optional, and when present is a bond, —O—,—C(═O)—, —S—, —S(═O)—, —S(═O)₂—, —NH—, —NHC(O)—, —C(O)NH—, —NR₉C(O)—,—C(O)NR₉—, —S(═O)₂NH—, —NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—, —OC(O)NH—,—NHC(O)O—, —OC(O)NR₉—, —NR₉C(O)O—, —CH═NO—, —ON═CH—, —NR₁₀C(O)NR₁₀—,heteroaryl-, aryl-, —NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—, —C(═NR₁₁)NR₁₀—,—OC(═NR₁₁)—, or —C(═NR₁₁)O—; L₄ is optional, and when present is a bond,substituted or unsubstituted alkyl, substituted or unsubstitutedcycloalkyl, substituted or unsubstituted alkenyl, substituted orunsubstituted alkynyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, substituted or unsubstituted heterocycle; orL₃, X and L₄ taken together form a nitrogen containing heterocyclicring; G is

wherein, R₆, R₇ and R₈ are independently selected from among H, halogen,CN, OH, substituted or unsubstituted alkyl or substituted orunsubstituted heteroalkyl or substituted or unsubstituted cycloalkyl,substituted or unsubstituted heterocycloalkyl, substituted orunsubstituted aryl, substituted or unsubstituted heteroaryl; each R₉ isindependently selected from among H, substituted or unsubstituted loweralkyl, and substituted or unsubstituted lower cycloalkyl; each R₁₀ isindependently H, substituted or unsubstituted lower alkyl, orsubstituted or unsubstituted lower cycloalkyl; or two R₁₀ groups cantogether form a 5-, 6-, 7-, or 8-membered heterocyclic ring; or R₁₀ andR₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclic ring; oreach R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof. 2.Use of a compound of Formula (A) with allogeneic hematopoietic stemcells and/or allogeneic T-cells for treating a patient for alleviationof a bone marrow mediated disease, with alleviation of consequentlydeveloped graft versus host disease (GVHD), wherein the compound ofFormula (A) has the structure:

wherein: A is N; R₁ is phenyl-O-phenyl or phenyl-S-phenyl; R₂ and R₃ areindependently H; R₄ is L₃-X-L₄-G, wherein, L₃ is optional, and whenpresent is a bond, optionally substituted or unsubstituted alkyl,optionally substituted or unsubstituted cycloalkyl, optionallysubstituted or unsubstituted alkenyl, optionally substituted orunsubstituted alkynyl; X is optional, and when present is a bond, —O—,—C(═O)—, —S—, —S(═O)—, —S(═O)₂—, —NH—, —NR₉—, —NHC(O)—, —C(O)NH—,—NR₉C(O)—, —C(O)NR₉—, —S(═O)₂NH—, —NHS(═O)₂—, —S(═O)₂NR₉—, —NR₉S(═O)₂—,—OC(O)NH—, —NHC(O)O—, —OC(O)NR₉—, —NR₉C(O)O—, —CH═NO—, —ON═CH—,—NR₁₀C(O)NR₁₀—, heteroaryl-, aryl-, —NR₁₀C(═NR₁₁)NR₁₀—, —NR₁₀C(═NR₁₁)—,—C(═NR₁₁)NR₁₀—, —OC(═NR₁₁)—, or —C(═NR₁₁)O—; L₄ is optional, and whenpresent is a bond, substituted or unsubstituted alkyl, substituted orunsubstituted cycloalkyl, substituted or unsubstituted alkenyl,substituted or unsubstituted alkynyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, substituted or unsubstitutedheterocycle; or L₃, X and L₄ taken together form a nitrogen containingheterocyclic ring; G is

wherein, R₆, R₇ and R₈ are independently selected from among H, halogen,CN, OH, substituted or unsubstituted alkyl or substituted orunsubstituted heteroalkyl or substituted or unsubstituted cycloalkyl,substituted or unsubstituted heterocycloalkyl, substituted orunsubstituted aryl, substituted or unsubstituted heteroaryl; each R₉ isindependently selected from among H, substituted or unsubstituted loweralkyl, and substituted or unsubstituted lower cycloalkyl; each R₁₀ isindependently H, substituted or unsubstituted lower alkyl, orsubstituted or unsubstituted lower cycloalkyl; or two R₁₀ groups cantogether form a 5-, 6-, 7-, or 8-membered heterocyclic ring; or R₁₀ andR₁₁ can together form a 5-, 6-, 7-, or 8-membered heterocyclic ring; oreach R₁₁ is independently selected from H or substituted orunsubstituted alkyl; or a pharmaceutically acceptable salt thereof, andis administered prior to, concurrently with, or following theadministration of the allogeneic hematopoietic stem cells and/orallogeneic T-cells.
 3. The use according to claim 1, wherein L₃, X andL₄ taken together form a nitrogen containing heterocyclic ring.
 4. Theuse according to claim 3, wherein the nitrogen containing heterocyclicring is a piperidine group.
 5. The use according to claim 1, wherein Gis


6. The use according to claim 1, wherein the compound of Formula (A) is1-[(3R)-3-[4-amino-3-(4-phenoxyphenyl)pyrazolo[3,4-d]pyrimidin-1-yl]piperidin-1-yl]prop-2-en-1-one(ibrutinib).
 7. The use of claim 1, wherein the GVHD is sclerodermatousGVHD.
 8. The use of claim 1, wherein the GVHD is steroid resistant GVHD.9. The use of claim 1, wherein the GVHD is sclerodermatous GVHD, steroidresistant GVHD, cyclosporin-resistant GVHD, refractory GVHD, oral GVHD,chronic oral GVHD, reticular oral GVHD, erosive GVHD, or ulcerative oralGVHD.
 10. The use of claim 1, wherein the patient has a relapsed orrefractory CLL.
 11. The use of claim 1, wherein the amount of thecompound of Formula (A) prevents or reduces GVHD while maintaining agraft-versus-leukemia (GVL) reaction effective to reduce or eliminatethe number of cancerous cells in the blood of the patient.
 12. The useof claim 1, wherein the cell transplantation is a hematopoietic celltransplantation.
 13. The use of claim 1, wherein the compound of Formula(A) is administered at a dosage of between about 0.1 mg/kg per day toabout 100 mg/kg per day.
 14. The use of claim 1, wherein the amount ofthe compound of Formula (A) is about 40 mg/day, about 140 mg/day, about280 mg/day, about 420 mg/day, about 560 mg/day, or about 840 mg/day. 15.The use of claim 1, wherein the compound of Formula (A) is administeredorally.
 16. The use of claim 1, wherein the compound of Formula (A) isadministered in combination with one or more additional therapeuticagents.
 17. The use of claim 16, wherein the additional therapeuticagent is a corticosteroid, cyclosporine (CSA), mycophenolate mofetil(MMF), or a combination thereof.